JP4790996B2 - Method for producing functional material from brewing by-products and functional material obtained thereby - Google Patents

Description

  The present invention relates to a method for producing a functional material from a brewing by-product and a functional material obtained thereby, and more specifically, koji or defatted koji or brown rice is treated with a hemicellulase enzyme, a liquefaction enzyme, and a saccharification enzyme. Next, the present invention relates to a method for producing a functional material characterized by treating with yeast and a functional material obtained thereby. The present invention further relates to a functional food, a pharmaceutical composition and a cosmetic composition containing the functional material.

  Most of the brewing by-products of beer and sake are currently treated as industrial waste. For example, red potato, a by-product of sake brewing, has been used as a feed at best, despite the fact that it contains abundant nutritional components, and until now no technology has been developed for effective use. Red cocoons are particularly rich in useful components such as L-arabinose, D-xylose, and various oligosaccharides (for example, arabinoxylan). These components are said to have various favorable effects such as dieting and health promotion. Conventionally, many methods for separating, purifying, or concentrating these components have been proposed (see Patent Documents 1 to 8).

  For example, Patent Document 1 relates to a method for purifying L-arabinose, and Patent Document 2 relates to a method for separating L-arabinose, both of which are characterized by the action of microorganisms that assimilate xylose. Patent Document 3 relates to a method for producing L-arabinose or an L-arabinose-containing enzyme-treated product and a method for producing L-arabinose, and Patent Document 4 describes a method for producing L-arabinose, an L-arabinose-containing enzyme-treated product, and a product thereof. Although it relates to the production method, none of them mentions D-xylose in the production method of L-arabinose in which arabinopolysaccharide is treated with an enzyme. Patent Document 5 relates to a method for producing water-soluble saccharides from cereal hulls, and relates to a method for producing L-arabinose, D-xylose and oligosaccharides thereof from arabino polysaccharides by acid treatment. Patent Document 6 relates to a method for preparing a water-soluble arabinoxylan, Patent Document 7 relates to an immunity enhancing substance and a method for producing the same, and Patent Document 8 describes a method for producing a water-soluble arabinoxylan from an arabino polysaccharide and the function thereof. It is about sex.

However, any prior art uses L-arabinose, D, which uses a brewing by-product such as the present invention, such as koji (red koji, defatted koji), or brown rice, as a raw material, and uses a two-step process of enzyme treatment and yeast treatment. -It does not relate to an efficient production technique of functional materials containing abundant oligosaccharide components such as xylose and arabinoxylan, nor does it suggest these.
JP 2002-262899 A Japanese Patent Laid-Open No. 2002-209597 JP 2002-95491 A JP 2001-286294 A Japanese Patent Laid-Open No. 11-113600 Patent No. 3079115 Japanese Patent No. 3347163 Japanese Patent Laid-Open No. 9-23895

  As described above, brewing by-products that are generated in large quantities but have no use or are rarely found are used as raw materials, are rich in L-arabinose, D-xylose and various oligosaccharides, and have favorable functions for health. It was also an object of the present invention to obtain a functional material having high safety and good fragrance.

  The inventors of the present invention have made extensive studies in view of the above circumstances, treating rice bran or defatted rice bran or brown rice with an enzyme containing hemicellulase enzyme and liquefaction enzyme and saccharifying enzyme as essential components, and then treating with yeast. The inventors have found that a functional material having a liver function improving function, an antiallergic function, an antioxidant function, a digestion absorption suppressing function, a blood sugar level increase suppressing function and the like can be obtained, and the present invention has been completed.

  According to the present invention, a functional material having a liver function improving function, an antiallergic function, an antioxidant function, a digestion absorption suppressing function, a blood sugar level increasing suppressing function, etc., and a high safety, and a functional food containing the same Pharmaceutical compositions as well as cosmetic compositions are obtained.

That is, the present invention
(1) A method for producing a functional material, comprising treating rice bran or brown rice with a hemicellulase enzyme and an enzyme system containing a liquefaction enzyme and a saccharification enzyme, and then treating with yeast.
(2) The method according to (1), wherein the cocoon is red cocoon;
(3) Functional material obtained by the method according to (1) or (2);
(4) a functional food, pharmaceutical composition or cosmetic composition containing the functional material according to (3); and (5) consisting of liver function improvement, antiallergy, antioxidant, digestion absorption suppression, and blood glucose level increase suppression The functional material according to (3) and the functional food or pharmaceutical composition according to (4), which have a function / efficacy selected from the group.

  As described above, the present invention provides a method for producing a functional material characterized in that koji or defatted koji or brown rice is treated with an enzyme system containing a hemicellulase enzyme and a liquefying enzyme and a saccharifying enzyme, and then treated with yeast. And a functional material obtained by the method. The functional material obtained is rich in L-arabinose, D-xylose and various oligosaccharides. Therefore, the raw material of the functional material of the present invention preferably has a high arabinoxylan content, and examples thereof include rice bran (red rice bran or defatted rice bran) or brown rice. A defatted lees is obtained by defatting red potatoes. A preferred raw material to be used in the present invention is red cocoon or defatted cocoon.

  According to the method of the present invention, the raw material is treated with a hemicellulase enzyme and an enzyme system containing a liquefying enzyme and a saccharifying enzyme. In the present specification, this treatment step is also referred to as “enzyme treatment” or “enzyme treatment step”. This process is aimed at breaking down polysaccharides and releasing glucose from starch.

  The “hemicellulase enzyme” may be any enzyme as long as it has hemicellulose-degrading activity, and may or may not be purified. The origin may be any. As the “hemicellulase-based enzyme”, for example, commercially available enzyme agents such as hemicellulase “Amano” 90 (manufactured by Amano Enzyme Co., Ltd.), hemicellulase M (manufactured by HBI Co., Ltd.) and the like can be used.

  The “liquefaction enzyme” may be any enzyme as long as it has an activity to liquefy starch, and may be purified or not purified. The origin may be any. As the “liquefying enzyme”, for example, commercially available enzyme agents such as Kokugen L (manufactured by Daiwa Kasei Co., Ltd.) and orientase AO40 (manufactured by HBI Corporation) can be used.

  The “saccharifying enzyme” may be any enzyme as long as it has an activity of saccharifying starch, and may or may not be purified. The origin may be any. As the “saccharifying enzyme”, for example, commercially available enzyme agents such as Gluc 100S (manufactured by Amano Enzyme Co., Ltd.), Glutase AD, and Glutase S (all manufactured by HTV Corporation) can be used.

  The “hemicellulase-based enzyme and the enzyme system containing a liquefying enzyme and a saccharifying enzyme” are the above-mentioned “hemicellulase-based enzyme”, “liquefying enzyme” and “saccharifying enzyme”, and may contain other enzymes. An enzyme mixture. Other enzymes may have an action of promoting the decomposition of raw materials or decomposing lipids and proteins in the raw materials. Moreover, what promotes the release | release of glucose from a raw material and gives a favorable flavor to the manufactured functional raw material may be sufficient. Examples of other enzymes include pectinase, mannanase, cellulase, α-galactosidase, phytase, arabanase, lipase, and protease. Depending on the purpose, these enzymes can be used singly or in combination. These enzymes may or may not be purified. Further, their origin may be any. As these enzymes, for example, pectinase XP-534 (manufactured by Nagase ChemteX Corporation), pectinase G “Amano” (manufactured by Amano Enzyme Co., Ltd.), cellulosin GM5 (mannanase) (manufactured by HI Corporation), lipase A-10FG ( Nagase ChemteX Corporation), Lipase AY “Amano” 30G (Amano Enzyme Co., Ltd.), Sumiteam PHY (Phitase) (Shin Nihon Chemical Co., Ltd.), Cellulosin TP25 (Xylanase) (manufactured by IB Corporation), etc. An enzyme agent may be used.

  The treatment conditions in the enzyme treatment step (for example, the amount of enzyme relative to the raw material, reaction temperature, pH, reaction time, etc.) can be appropriately selected according to the purpose. Examples of the ratio (weight ratio) of the enzyme to the raw material are hemicellulase enzyme 1/20 to 1/50, liquefaction enzyme 1/50, saccharification enzyme 1/100, lipase, and phytase 1/500. If the amount of the enzyme is too small, the reaction is insufficient, or the reaction takes a long time. If the amount of the enzyme is too large, it is not economically preferable. The reaction temperature should be determined in consideration of the optimum temperature of the enzyme used, for example, 15 to 60 ° C. will generally be used. The reaction pH should also be determined in view of the optimum pH of the enzyme used and will generally be in the range of 3-10. As shown in the examples, a pH adjusting buffer may not be used. The reaction time can be determined as appropriate, but if it is too short, the reaction is insufficient, and if it is too long, problems such as contamination with germs occur.

  The enzyme-treated product obtained by the enzyme treatment step is treated with yeast. In the present specification, this treatment step is also referred to as “fermentation”, “yeast treatment”, “fermentation step” or “yeast treatment step”. The characteristic of this yeast process is that pentose (L-arabinose, D-xylose, etc.) is a non-fermentative (utilizing) glucose, fructose fermenting (utilizing) sake yeast that concentrates pentose and produces microbial microorganisms It is to prevent contamination and to proliferate yeast to obtain many yeast cells. Regarding yeast cells, it has been found that dietary fibers derived from cell wall components have abundant functions, and the yeast cells themselves contain various nutrients and are considered good for the gastrointestinal tract. . Furthermore, preferable fermented aroma is produced by fermenting with yeast.

Although the reaction conditions (reaction temperature, pH, time, used yeast, aeration conditions, etc.) in the yeast treatment step can be appropriately determined, the conditions should be suitable for the characteristics of the yeast used such as glucose consumption and growth. Examples of yeast used include Saccharomyces genus, Schizosaccharomyces genus, Candida genus, Kluyveromyces genus, etc., considering the use for functional foods and pharmaceutical compositions, Saccharomyces genus yeast is preferred, especially sake yeast, Beer yeast is preferred. The amount of yeast inoculation can also be selected as appropriate, but will generally be on the order of 10 ⁷ to 10 ⁸ cells / ml. Ventilation can be performed as needed.

  The functional material obtained by the production method of the present invention described above contains a large amount of oligosaccharides such as L-arabinose and D-xylose and arabinoxylan, and also contains a large amount of yeast cells, which will be described in detail below. It has abundant functions. As the functional material of the present invention, the processed yeast product obtained by the above method may be used as it is, or it may be used after further processing. For example, it may be further concentrated or diluted, may be used after being subjected to a drying treatment such as freeze-drying or heat-drying, or may further be used after extracting a solid content with water or the like. The form is not particularly limited, and may be, for example, a solution, a suspension, a semi-solid (for example, a paste), a solid (for example, a powder or a granule), and the like. As will be described in Examples, the functional material of the present invention exhibits various activities / functions that are considered to be good for health, and typically has a liver function improving function, an antiallergic function, an antioxidant function, and digestive absorption. Various physiological activities / functions and pharmacological activities / functions such as an inhibitory function and a blood glucose level elevation inhibiting function are shown. In addition, the functional material of the present invention is highly safe and can be used for foods and pharmaceuticals. Here, “functional food (material)” means “nutrient function (primary function) inherent to food (material), sensory function (secondary function) such as taste and aroma, Pay attention to biological regulation functions (tertiary functions) such as biological defense, prevention and recovery of diseases, adjustment of physical rhythm, suppression of aging, etc., so that these biological regulation can be elucidated scientifically and functions can be demonstrated Means food (material) that has been designed and processed.

  The functional material obtained by the present invention can be used for various applications. The functional material of the present invention has a good scent. As one use, various functional foods or health foods can be obtained by using the functional material of the present invention in foods. For example, it may be used as it is or in the form of pellets, powders, granules or the like, or may be used as a food additive, seasoning or sprinkle. Moreover, you may use the functional raw material of this invention, making it contain in a foodstuff.

  As a further use, the functional material of the present invention can be used in a pharmaceutical composition. In general, such pharmaceutical compositions will include a pharmaceutical carrier or excipient in addition to the functional material of the present invention. The form is not particularly limited, and for example, it may be solid (tablet, powder, granule, etc.), semi-solid (pasta etc.), liquid (solution, suspension, emulsion, etc.), or encapsulated in a capsule. Good. Carriers or excipients for these dosage forms are also well known to those skilled in the art and can be selected as appropriate.

  As another application, the functional material of the present invention can be used in a cosmetic composition. Since the functional material of the present invention is rich in ferulic acid and contains little or no glucose, there is no stickiness to the skin by glucose, and there is an active oxygen scavenging effect and a skin cell growth promoting effect by ferulic acid, There are uses as cosmetic materials. Therefore, the present invention relates to a cosmetic composition containing the functional material of the present invention. The cosmetic composition of the present invention contains a carrier commonly used in cosmetics in addition to the functional material of the present invention, such as water, glycerin, alcohols, oils and fats, and further includes a fragrance, a colorant and a preservative. Etc. may be added as appropriate. The shape of the cosmetic composition containing the functional material of the present invention is not particularly limited, and for example, it can be a lotion, emulsion, cream, ointment or the like.

  These functional foods and pharmaceutical compositions have the functions / effects of the functional material of the present invention, and typical functions / effects thereof include, for example, liver function improvement, anti-allergy, anti-allergy, Oxidation, digestion absorption inhibition, and blood glucose level elevation inhibition are mentioned.

  The present invention will be described in more detail and specifically with reference to the following examples, but these examples should not be construed as limiting the present invention.

Screening of enzymes used for enzyme treatment (1)
Dissolve 2 g of wheat-derived arabinoxylan (Megazyme) in 10 ml of 50 mM phosphate buffer (pH 5.0), hemicellulase agent, pectinase agent, xylanase agent, cellulase agent, phytase, lipase, α -L-arabinose (hereinafter also referred to as “arabinose”), D-xylose (hereinafter referred to as “the arabinose”) after adding 100 mg each of commercially available enzyme agents such as galactosidase, mannanase, arabanase, α-amylase, glucoamylase and reacting at 50 ° C. for 18 hours. The enzyme agent from which arabinoxylan oligosaccharides are released was also screened. Qualitative analysis of oligosaccharides of arabinose, xylose, and arabinoxylan was developed with thin layer chromatography (TLC plastic sheet Sirica gel 60 F254 (Merck), developing solvent ethyl acetate: pyridine: water = 12: 5: 4, concentrated sulfuric acid. Quantitative analysis was performed with liquid chromatography (HPLC system (Gilson), column Asahipak NH2P-50 4E (Showa Denko), SEDEX-55 evaporative light scattering detector (SEDERE)).

As a result, the enzyme agents that release a large amount of arabinose are Sumiteam ARS, Sumiteam PX (Shin Nippon Chemical Industry Co., Ltd.), Hemicellulase “Amano” 90, Cellulase A “Amano” 3 (Amano Enzyme Co., Ltd.), Cellulosin HC100 ( Enzyme agents that release a large amount of xylose are pectinase G “Amano”, hemicellulase “Amano” 90 (manufactured by Amano Enzyme), cellulosin TP25, cellulosin GM5, cellulosin T2 (manufactured by HBI Corporation) ), Sumiteam ACH (manufactured by Shin Nippon Chemical Industry Co., Ltd.), enzyme agents that release a large amount of arabinoxylan oligosaccharides are pectinase XP-534 (manufactured by Nagase ChemteX Corporation), VERON191 (manufactured by Rame Enzyme), etc. I understood.
Next, as a result of examining conditions under which the amount of monosaccharides of arabinose and xylose is high and conditions of a large amount of oligosaccharides of arabinoxylan by combining various enzyme agents under the same reaction conditions using red koji and defatted lees In red rice bran, it was found that monosaccharides were released efficiently when pectinase XP-534 and cellulosin GM5 were combined, and in defatted silkworms, Sumiteam ARS and cellulosin GM5 were combined.

  In addition, lipase A-10FG (manufactured by Nagase ChemteX Corporation), Sumiteam PHY (Nippon Chemical Industry Co., Ltd.) are enzyme enzymes that simultaneously treat lipase and phytase during the red yeast enzyme reaction to improve the taste without bitterness. Selected). Furthermore, as an enzyme agent that efficiently degrades starch from red koji and defatted koji to glucose, liquefied amylase agent Kokugen L (manufactured by Yamato Kasei Co., Ltd.) and saccharified amylase agent gluc 100S (manufactured by Amano Enzyme Co., Ltd.) It turned out to be a good combination.

Preparation of functional materials 1
In response to the results of the enzyme treatment conditions in the preliminary test, red candy and defatted candy were subjected to enzyme treatment under the following conditions. In addition, the powder dried product of the functional material obtained from the red koji according to this example is referred to as “red koji fermented product 1”, and the powder dried product of the functional material obtained from the degreased koji is referred to as “fat koji fermented product 1 "

  Add 10kg of red potato, 50L of water, 500g of pectinase XP-534, 20g of lipase A-10FG, pH 7, 18 hours at 50 ° C, 500g of cellulosin GM5, 20g of Sumiteam PHY, 200g of Kokugen L, 100g of Gluc 100S In addition, the enzyme treatment was performed at pH 4.5 and 55 ° C. for 18 hours. Add 10kg of defatted rice cake, 50L of water and 500g of Sumiteam ARS, pH 3.5, 18 hours at 55 ° C, add 500g of cellulosin GM5, 200g of Kokugen L, 100g of Gluc 100S, 18 hours at pH 4.5, 55 ° C Processed. Each pH adjustment was performed with lactic acid for food addition and sodium lactate. The glucose and fructose content after the enzyme treatment was measured (in the column before fermentation in Table 1).

Inoculate 1 × 10 ⁸ froth-free yeast B29, a sake yeast, into the red and defatted enzyme treatment solution, further increase the number of yeast after fermentation (enhanced various functions of sake yeast), In order to suppress the propagation of various bacteria, 5 kg of glucose was added, and stirring culture was performed at 25 ° C. for 5 days. Yeast count after fermentation (cells / ml), alcohol concentration (Alc;%), glucose level (Glu;%), fructose level (Fru;%) and titrated acidity (TA; ml), titrated amino acid level (AA; ml) Table 1 shows a comparison of analytical values before and after fermentation.

  From Table 1, the number of yeast increases in both red koji fermented liquor and defatted koji fermented liquid, and glucose and fructose are almost assimilated by sake yeast and converted to alcohol. There is no change in xylose before and after yeast fermentation, the starch content of saccharides is reduced, arabinose and xylose are concentrated, the number of yeasts is increased, and the material that achieves the purpose of preventing microbial contamination with the generated alcohol has been developed did it. The titrated acidity of the defatted koji fermented liquid was higher than that of the red koji fermented liquid, and both became a slightly sour material, but both had good aroma after fermentation.

  Powder drying is performed by heating and concentrating each fermented liquid twice (100 ° C, 5 hours) and using a spray dryer SD-1000 (manufactured by Tokyo Rika Kikai Co., Ltd.). 1.25 kg of 1 was obtained. Table 2 shows the general component values of the dried product.

  Compared with the rice bran component of the food composition table from Table 2, all fermented products had a higher number of yeasts, and the content of dietary fiber and protein derived from the yeast cell wall component and yeast cytoplasm component increased. In both cases, the starch content was reduced due to the degradation of starch. In red koji fermented product 1, the degradation of lipid became part due to the treatment with the lipase agent.

  The free arabinose and xylose content of each fermented product and the content of arabinose and xylose polysaccharide remaining in the insoluble residue were 72% sulfuric acid, stirred at room temperature for 1 hour, then 4% sulfuric acid, 121 ° C autoclaved for 1 hour. Table 3 shows the results of measurement of the arabinose and xylose contents after the decomposition.

  From Table 3, the percentages of soluble (free / free + residue) arabinose and xylose in red koji fermented product 1 are 12% and 4%, respectively, and the proportions of soluble arabinose and xylose in defatted koji fermented product 1 are 13% and 3%, respectively. It became. Both the red koji fermented product 1 and the defatted koji fermented product 1 were found to contain soluble arabinoxylan oligosaccharides that were not assimilated by yeast fermentation after enzyme treatment in thin-layer chromatograms and liquid chromatograms. Could not (because there is no reference material).

Screening of enzymes used for enzyme treatment 2
Commercially available hemicellulase agent, pectinase agent, xylanase agent, cellulase agent, phytase, lipase, α-galactosidase, mannanase, arabanase, etc., which are commercially available food enzymes, in 2 g of Nihonbare rice produced in 2003 and 10 ml of 50 mM phosphate buffer 100 mg of each enzyme agent was added, and after reacting for 18 hours at the optimum pH and temperature of each enzyme agent, the turbidity before and after the reaction (HACH, 2100N Turbidimeter), glucose amount, and direct reducing sugar amount were measured. As a result, Sumiteam ARS, Sumiteam PX, Sumiteam ACH (manufactured by Nippon Chemical Industry Co., Ltd.), pectinase G “Amano”, hemicellulase “Amano” 90 (manufactured by Amano Enzyme Co., Ltd.), pectinase XP-534 (Nagase ChemteX Corporation) It was found that enzyme agents such as Cellulosin TP-25, Cellulosin GM5 (manufactured by HIBI Co., Ltd.) and VERON191 (manufactured by Rame Enzyme) have high turbidity, glucose amount and direct reducing sugar increase rate. Above all, the rate of increase in turbidity of cellulosin TP-25 was high, resulting in a strong enzyme that acts on the outermost part of brown rice.

Next, red cocoon, defatted cocoon 10 g each and 50 mM phosphate buffer 50 ml, Sumiteam ARS, Sumiteam PX, Sumiteam ACH, pectinase G `` Amano '', hemicellulase `` Amano '' 90, pectinase XP-534, cellulosin TP-25, Add 500 mg of cellulosin GM5 and VERON191 enzyme agents, react for 18 hours at optimum pH and temperature of each enzyme agent, inoculate 5 × 10 ⁷ sake yeast B29, add 5 g of glucose at 25 ° C Fermentation was carried out for 4 days, and free arabinose, xylose content and titratable acidity were measured. As a result, it was found that pectinase G “Amano” and hemicellulase “Amano” 90 independently release a large amount of arabinose and xylose, and the titrated acidity of the fermentation broth can be lowered.

Furthermore, 500 mg of 3 types of enzyme agents, Cellulosin TP-25, Pectinase G “Amano”, and Hemicellulase “Amano” 90, were added to 10 g each of red koji and defatted koji and 50 mM phosphate buffer, changing the order of combination. After each reaction for 18 hours at the optimum pH and temperature of each enzyme agent, inoculate 5 × 10 ⁷ sake yeast B29, add 5 g of glucose, ferment at 25 ° C. for 4 days, and release arabinose, The xylose content and titratable acidity were measured. As a result, both red koji and defatted koji fermented liquor were reacted with cellulosin TP-25 first, and then pectinase G “Amano” and hemicellulase “Amano” 90 were reacted simultaneously to release a large amount of arabinose and xylose. It was found that the titratable acidity of the fermentation broth can be lowered.

In addition, 10 g each of red koji and defatted koji and 50 ml of 50 mM phosphate buffer, 100, 200 and 500 mg of Kelogen L, 3 types of enzyme agents, cellulosin TP-25, pectinase G “Amano” and hemicellulase “Amano” 90, respectively. 200 mg and 100 mg of Gluc 100S, and in Akamine, lipase AY “Amano” 30G (manufactured by Amano Enzyme Co., Ltd.) or lipase A-10FG 20 mg each and Sumiteam PHY 20 mg were added, and the optimum pH of each enzyme was After reaction for 18 hours or 42 hours at appropriate temperature, inoculate 5 × 10 ⁷ sake yeast B29, add 5g of glucose, ferment at 25 ° C for 4 days, and measure free arabinose, xylose content and titration acidity did. As a result, in red sea bream, 200-500 mg of cellulosin TP-25, 200 mg of Kokugen L and 100 mg of glucogen 100S were reacted for 18 hours at pH 5 and 60 ° C., then pectinase G `` Amano '' and hemicellulase `` Amano '' 90 were 200-200 After reacting 500 mg and Sumiteam PHY at 20 mg, pH 4.5, 50 ° C. for 18-42 hours, reacting lipase AY “Amano” 30G at 20 mg, pH 7, 50 ° C. for 18 hours, and fermenting arabinose and xylose after fermentation in yeast It was found that the conditions were free. On the other hand, in defatted lees, 100-200 mg of cellulosin TP-25, 200 mg of kokugen L and 100 mg of glucogen 100S at pH 5 and 18 hours at 60 ° C., then pectinase G “Amano” and hemicellulase “Amano” 90 100-200 mg The reaction was carried out at pH 4.5 and 50 ° C. for 18 to 42 hours, and it was found that the conditions were such that a large amount of arabinose and xylose after fermentation was released in yeast. Moreover, the arabinose, xylose, and titratable acidity which a red koji fermented liquid and a defatted koji fermented liquid liberated were also comparable.

Preparation of functional materials 2
In response to the results of the enzyme treatment and fermentation conditions in the preliminary test, the enzyme treatment and fermentation of red koji and defatted koji were performed under the following conditions. In addition, the freeze-dried product of the functional material obtained from red koji according to this example is referred to as “red koji fermented product 2”.

After reaction with 2 kg of red potato, 10 g of water and 40 g of cellulosin TP-25 or 100 g and 40 g of Kokugen L and 20 g of Gluc 100S at pH 5 and 60 ° C. for 18 hours, 40 g of pectinase G “Amano” and hemicellulase “Amano” 90 Or after reacting 100g and Sumiteam PHY 4g at pH 4.5, 50 ° C for 42 hours (category with low amount of enzyme) or 18 hours (category with high amount of enzyme), 4g of lipase AY "Amano" 30G The reaction was carried out at pH 7, 50 ° C. for 18 hours. After reacting 2 kg of defatted water, 10 L of water, 40 g of cellulosin TP-25, 40 g of Kokugen L and 20 g of Glucogen 100S at pH 5 and 60 ° C. for 18 hours, pH 4 at 40 g of pectinase G “Amano” and hemicellulase “Amano” 90 The reaction was carried out at 50 ° C. for 42 hours. The pH was adjusted with hydrochloric acid and sodium hydroxide for food addition. Inoculate 5 x 10 ⁷ sake yeast B29 into 2 sections of red koji enzyme treatment solution and 1 section of defatted koji enzyme treatment solution, add 1 kg of glucose and perform stirring fermentation at 25 ° C for 4 days, free arabinose, xylose, Glucose content, yeast number, alcohol concentration, titratable acidity and titrated amino acid degree were measured (Table 4). As a result, the analysis values of the two sections of the red koji fermentation liquid and the one section of the defatted koji fermentation liquid were almost the same. Therefore, the fermented material with a small amount of the enzyme enzyme of red koji was directly dried with a vacuum freeze dryer TF10-80TPA (manufactured by Takara Manufacturing Co., Ltd.) without heat treatment to obtain 1.66 kg of red koji fermented material 2. . Table 4 shows a comparison of various components of the red koji fermented product 2 and the raw red koji fermented product 1.

  According to Table 4, fermented red koji 2 had red koji soluble (free / free + residue) arabinose and xylose ratios of 35% and 23%, respectively. In addition, compared to fermented red koji 1, a material that was 3 times more free arabinose and 6 times more free xylose could be developed. The oligosaccharides of soluble arabinoxylan of this red koji fermented product 2 also existed as oligosaccharides not assimilated by yeast fermentation after enzyme treatment in the liquid chromatogram, but could not be quantified. In addition, the number of yeast in this red koji fermentation broth (before freeze-drying red koji fermented product 2) was about the same as the previous one, but the production of alcohol was slightly higher and glucose was completely lost. The titratable acidity can be lowered to about 40% of the previous time, and it has become a material with excellent fertility and taste that does not stand out sourness.

in vitro test
(A) Disaccharide-degrading enzyme inhibition test In order to evaluate the digestion and absorption inhibition effect of the obtained red koji fermented product and defatted koji fermented product, the maltase activity inhibition rate, sucrase activity using the small intestine crude enzyme which shows the preparation method The inhibition rate was measured.
Rat intestinal acetone powder (manufactured by Sigma) was added with 10 volumes of maleic acid buffer (pH 6.0), centrifuged at 5000 rpm for 30 minutes, and the resulting supernatant was used as a small intestine crude enzyme. In addition, after adding 3.33 volumes of maleic acid buffer (pH 6.0) to the powder sample and centrifuging at 14000 rpm for 20 minutes, the supernatant obtained was evaluated by inactivating the residual enzyme in a 70 ° C hot water bath. did.

The maltase activity inhibition rate is the mixture of the small intestine crude enzyme and the extract in the final concentration 1% maltase solution. The sucrase activity inhibition rate is the mixture of the small intestine crude enzyme and the extract in the final concentration 1% sucrose solution, each at 37 ° C for 1 hour. The amount of glucose produced in the solution after the reaction was measured with a glucose cII test Wako (manufactured by Wako Pure Chemical Industries, Ltd.). Moreover, the inhibition rate was derived from the following calculation formula based on the blank value without mixing the extract. The results are shown in Table 5.
Activity inhibition rate (%) = 100−B / A × 100
A: Blank glucose production, B: Extract glucose production

  From Table 5, both the red koji fermented product (both 1 and 2) and the defatted koji fermented product 1 were found to have sucrase activity inhibition and maltase activity inhibition effects. It is considered that glucose is reduced in these fermented products, and oligosaccharides of arabinose, xylose, and arabinoxylan are concentrated. There was a tendency for both activities to be increased due to the influence of the coexisting sugars in the raw red koji and defatted koji.

(B) DPPH radical scavenging ability test In order to evaluate the antioxidant capacity of fermented red koji and fermented koji koji, 20 times volume of 50% EtOH was added to the powder sample, extracted at room temperature for 3 hours, 3000 rpm, 5 minutes After centrifugation, the obtained supernatant was subjected to DPPH radical scavenging ability test.

25 μL of 2M acetate buffer (pH 5.5), 475 μL of 99.5% EtOH, 500 μL of extract, and 125 μL of 1,1-Diphenyl-2-picrylhydrazyl (DPPH) were mixed, and OD517nm was measured 30 minutes after addition of DPPH. In order to calculate radical scavenging ability, water mixed in place of the specimen was also measured.
The radical scavenging ability was calculated from the following formula for radical scavenging ability.
Radical scavenging ability (%) = 100− (OD517 _B / OD517 _A ) × 100
OD517 _A : Water (control), OD517 _B : Specimen Glutathione, which is known to have high DPPH radical scavenging ability, was used as a standard, and the radical scavenging ability of the fermented product and the control was converted to the amount of glutathione. The results are shown in Table 6.

  From Table 6, both the red koji fermented product (both 1 and 2) and the defatted koji fermented product 1 were found to have DPPH radical scavenging ability, that is, antioxidant ability. Both raw red koji and defatted koji have antioxidant capacity, but it has been found that fermenting has the effect of increasing their activity.

Safety test
(A) Acute oral toxicity screening test To examine the acute oral toxicity of fermented red koji fermented product 1 and defatted koji fermented product 1 to rats, three female (6 weeks old) Slc: Wistar (SPF) animals were used. The test was conducted. The dose was 1 dose of 2000 mg / kg. After weighing precisely 2000 mg of fermented red koji fermented product 1 and defatted koji fermented product 1, a suspension of 10 mL of distilled water added for injection was given to the animal fasted for 16 hours before administration using a stomach tube. The observation period after administration was 7 days, and the animals were observed for life and death, general condition, and body weight change, and pathological anatomical examination was performed at the end of the observation (Table 7).

(B) Reverse mutation screening test In order to examine the gene mutagenicity of fermented red koji fermented product 1 and defatted koji fermented product 1, a reverse mutation screening test using Salmonella typhimurium TA100 and TA98 strains was conducted. It was.

  The test substance was prepared by dissolving or suspending in a suitable solvent to prepare a preparation stock solution, and sterilizing by filtration if necessary. As test doses, 8 doses (common ratio 2) of 39.1, 78.1, 156, 313, 625, 1250, 2500 and 5000 μg / plate were used. In order to confirm the growth inhibitory action of the test substance, the growth state of the test strain on the plate was observed with an actual microscope, and the deposition state of the test substance was further observed with the naked eye. Next, the number of colonies generated by back mutation was counted, and when the number of back mutation colonies increased more than twice that of the negative control and the increase was confirmed to be dose-dependent, it was determined to be positive. (Table 7).

(C) Residual pesticide multi-component simultaneous analysis According to the conventional method, after pretreatment such as extraction, concentration, column purification, etc. of red koji fermented product 1, screening analysis of 77 pesticide residues was performed by GC-MS measurement. (Table 7).

(D) Heavy metal analysis The heavy metal in the red yeast rice fermented material 1 was measured with the following method.
Arsenic, lead and cadmium were measured by atomic absorption spectrophotometry, total mercury was measured by reductive vapor atomic absorption spectrophotometry, total chromium was measured by diphenylcarbazide spectrophotometry, and cyan by pyridinepyrazolone spectrophotometry (Table 7).

  From Table 7, both the red koji fermented product 1 and the defatted koji fermented product 1 were negative in both the acute oral toxicity screening test and the reverse mutation screening test. As a result of simultaneous analysis of 77 pesticide residues, 77 components were below the limit of quantification. The results of heavy metal analysis of arsenic, lead, mercury, cadmium, cyan, and chromium were not problematic. That is, it was found that both the red koji fermented product 1 and the defatted koji fermented product 1 have no problem in safety as food materials. Similar results were obtained for fermented red koji 2. Therefore, the safety of the functional material of the present invention was confirmed.

Blood glucose level increase inhibitory effect confirmation test
4-week-old Sprague-Dawley male rats (Jc1; SD, manufactured by Clea Japan Co., Ltd.), room temperature 23 ± 1 ° C, humidity 55 ± 7%, light / dark cycle 12 hours (light period 8-20 hours), solid MF Feed (Oriental Yeast Co., Ltd.) and water were freely given and subjected to the experiment after preliminary breeding for 4 weeks (8 weeks old, 264.6 ± 5.1 g). Unanesthetized and unrestrained rats with catheters placed in the stomach and portal vein were divided into 2 groups of 5 each and then reared for 24 hours, then fasted for 16 hours, then 15% sucrose solution 11.25 ml / kg rat / hour It was confirmed that the glucose concentration in the portal vein blood became constant after administration for 2 hours at a rate of 5%. Extract the red koji fermented product 2 and the control red koji for 30 minutes each with 3 times the amount of water, centrifuge at 14000 rpm for 20 minutes, filter through 250 mesh, and filter with the No. 5A filter paper. Each concentrated sample was administered into the stomach (red bean ferment extract; 0.36 g / kg arabinose + xylose equivalent, red bean; similar operation control), then portal catheter up to 3 hours every 10 minutes Further, 0.05 ml of portal vein blood was collected, and the glucose concentration in the plasma was measured (FIG. 1; rat administered with red yeast ferment extract extract, FIG. 2; rat administered with control red coral).

1 and 2, the glucose concentration in the portal vein blood was significantly decreased 20 minutes after administration in the red yeast ferment extract (indicated by an asterisk in FIG. 1). In the control red rabbit, the glucose concentration in the portal vein blood was not significantly decreased, but was significantly increased at 10 minutes and 100 minutes after administration. In the total concentration of arabinose and xylose, 0.36 g / kg rat was found to have an inhibitory effect on α-glucosidase in the rat small intestine. Therefore, it was found that the fermented red koji product of the present invention is a material capable of suppressing hyperglycemia after diabetic diet.

Liver function improvement effect confirmation test
6 weeks old SD male rats (CLEA Japan, Inc.) were preliminarily raised for 1 week, then feed and water were freely consumed with the diet shown in Table 8, room temperature 23 ± 2 ° C, humidity 60 ± 10%, 12 Breeding experiments were conducted for 20 days in an environment with a light-dark cycle.

  On day 21 of breeding, liver and cecal weights, total cholesterol and triglycerides (TG) for tail vein serum, GOT (glutamic oxaloacetic transaminase activity) and GPT (glutamic pyrvic transaminase activity) for abdominal aorta plasma ), Glucose and HDL-cholesterol (HDLcho) were measured (Table 9).

  From Table 9, the fermented red koji fermented product 1 (Group C) and defatted koji fermented product 1 (Group D) have significantly reduced enzyme activities of GOT and GPT in plasma compared to cellulose (Group A). Both showed improvement of liver function. The red koji fermented product 1 has a tendency to slightly increase HDL-cholesterol, the cecal weight per body weight is also significantly increased, and it is considered that there is some dietary fiber effect.

Allergy reduction effect confirmation test
Alleviate Brown ssn slc rats at 4 weeks of age (4 from 1 group), using 4 groups as one group, MF solid feed and water ad libitum, and each feed (water, red potato, 4 groups of fermented red koji and fermented koji koji were administered with a gastric sonde to a dose of 60 mg, and the temperature was 23 ± 2 ° C, humidity was 60 ± 10%, and the environment was 12 hours light-dark cycle for 6 weeks (4 Breeding experiments were conducted between 10 weeks of age and 10 weeks of age. Plasma IgA, IgE, sIgA in feces and spleen and liver weights at 10 weeks of age were measured every week. As a result, body weight increased steadily in each administration group. In addition, there was no difference in spleen, liver weight, and fecal sIgA among the administration groups. 3 and 4 show changes in plasma IgA in each administration group, and FIG. 5 shows changes in IgE.

  From FIG. 3, it can be seen that the plasma IgA value increased in the group administered with red yeast fermented product 1 after the 3rd week, and particularly increased in the 5th week. There was no difference in the other groups. Further, FIG. 4 shows that in the group administered with fermented red koji, the plasma IgA level significantly increased after the third week of administration, and the humoral immune mechanism was activated. Furthermore, from FIG. 5, in the red koji fermented product 2, a decrease in plasma IgE was significantly observed at 10 weeks of age, and the immune mechanism was activated. Therefore, it was found that fermented red koji 2 has an immunostimulatory function. Therefore, it was confirmed that the red koji fermented product of the present invention has an allergy reducing effect in allergic rats.

  The present invention provides functional materials having various functions preferable for health and high safety, and can be used in the fields of foods, medicines, and cosmetics.

It is a figure which shows the time-dependent change of the glucose level in the plasma in a red yeast fermented extract extract administration group. * Indicates significance at 5% risk. It is a figure which shows the time-dependent change of the glucose level in the plasma in a control red rabbit administration group. FIG. 3 is a graph showing changes in blood IgA over time in four groups: a water administration group, a red koji administration group, a red koji fermented product 1 administration group, and a defatted koji fermented product 1 administration group. FIG. 4 is a diagram showing changes in IgA in the group administered with red koji fermented product 1. * Indicates significance at 5% risk. FIG. 5 is a view showing changes in IgE in plasma of the group administered with red yeast fermented product 2. * Indicates a significance level of 5%, and ** indicates a significance level of 1%.

Claims (3)

A method for producing a functional material, comprising treating red koji or defatted koji with an enzyme system containing a hemicellulase enzyme and a liquefying enzyme and a saccharifying enzyme, followed by fermentation with yeast, wherein the material is L-arabinose , D-xylose, and arabinoxylan in total 5% or more . A functional material obtained by the method according to claim 1 . A functional food, pharmaceutical composition or cosmetic composition comprising the functional material according to claim 2 .

Images