KR20200080442A - Concentrated liquid of glycosylated Lentinus edodes, Concentrated powder of glycosylated Lentinus edodes and Manufactured method of the same - Google Patents

Abstract

The present invention relates to a shiitake mushroom processed product and a manufacturing method thereof. More specifically, the present invention relates to a shiitake mushroom saccharification concentrate including a high content of amino acids, shiitake mushroom saccharification concentrate powder, and the manufacturing method thereof. According to the manufacturing method of the shiitake mushroom saccharification concentrate, after a concentrate product in which an extract product of shiitake mushroom dried powder is concentrated in saccharified with a saccharification enzyme, a saccharified product is re-concentrated.

Description

Shiitake mushroom saccharified concentrate, shiitake mushroom saccharified concentrated powder using the same, and its manufacturing method{Concentrated liquid of glycosylated Lentinus edodes, Concentrated powder of glycosylated Lentinus edodes and Manufactured method of the same}

The present invention relates to a shiitake mushroom saccharified concentrate rich in beneficial amino acids, a shiitake mushroom saccharified concentrate, and a method for preparing them.

Mushrooms contain sugars, lipids, proteins, minerals and nutrients such as vitamins evenly and have a unique taste and aroma, and have been widely used for food since ancient times. Recently, they have been spotlighted as non-polluting natural foods.

In addition, basidiomycetes contain various medicinal ingredients and have been used as a folk remedy since ancient times. So, in recent years, more interest in the components of the mushroom, which is a high-potency fungi, has been actively studied in the food and pharmacological aspects, and the nutritional and medicinal components of various mushrooms have been revealed. Among these components, proteins and amino acids are nutrients that have a great influence on biological function and metabolism, and are also closely related to flavor. Therefore, these ingredients have attracted the attention of many scholars.

Mushrooms are higher fungi that form fruiting bodies among basidiomycetes belonging to the fungus, and about 15,000 species are known worldwide. Of these, about 970 species are recorded in Korea, and about 20 are cultivated. Mushrooms have recently received a lot of attention as a functional food along with the pharmaceutical field because of their pharmacological effect. Mushrooms are an important factor that has attracted attention as medicines and new food materials, and they have the advantage that side effects are rarely exhibited even when taken for a long time for food or medical purposes. Mushrooms contain various nutrients that are important to the human body, such as vegetable proteins, amino acids, enzymes, fats, iron, fiber, vitamins, minerals, etc., and have been eaten by people for a long time because of their unique aroma and taste. It is known as a low-calorie food rich in protein. In addition, it is a food ingredient that contains a lot of various health functional substances, and has antioxidant activity for preventing aging, anti-cancer, anti-diabetes, weight loss, immunity enhancement, hypertension prevention, etc. It is widely used as a raw material for pharmaceuticals, and products for improving liver function and immunity using mycelium as well as fruiting bodies are commercially available. Recently, as mushrooms are sufficiently recognized by consumers as healthy foods, they have reached the stage of securing a fixed consumer base. Until now, most of the processes using basidiomycetes as food and pharmaceutical products mainly consisted of fruiting bodies or simple products in which these mycelium were liquid-cultured. Mushrooms are one of the top 10 health foods in the world, and consumption is increasing in proportion to the degree of economic development. Developed countries eat more than 5 kg per person per year. It is estimated that Korea consumes about 3kg, and consumption is expected to continue to increase as it is recognized as a low-calorie functional food suitable for the well-being era.

Shiitake mushrooms, which are easy to obtain in the surroundings and cheap, often come to our table. In'Donguibogam' and'Bonchogangmok', it is written that'it strengthens the air and does not feel hunger, thereby improving the wind and helping blood circulation'. It also has the effect of clearing blood and boosting appetite. When cooking pork, put it together for better absorption. The unique umami of shiitake mushrooms is that guanilic acid is more common than other mushrooms. Guanilic acid has the property of lowering cholesterol, which is good for people with high blood pressure and heart disease. In addition, lentinan in shiitake mushrooms is a powerful anti-cancer agent that activates the immune system. Therefore, it is effective not only for cancer, but also for viral diseases such as cold, high blood pressure, and diabetes. In addition, shiitake mushrooms are rich in various minerals and vitamins, and fiber helps digestion of the stomach and small intestine, and is good for obesity, diabetes, heart disease, and liver disease. In addition, it is rich in protein, calcium, phosphorus, and iron and is rich in ingredients such as vitamin D, which strengthens bones, vitamin B, which is essential for hematopoietic action, and eliteatine, which helps metabolize blood, so it is good for children in growing season. Sun-dried shiitake mushrooms are twice as nutritious as raw shiitake mushrooms. In particular, they are known to make them strong and prevent osteoporosis because they contain a lot of vitamin D to help absorb calcium.

In Korea, although wild and artificially grown mushrooms are widely edible, systematic research on the composition of edible mushrooms from the food side is insufficient, and studies on the lipids of oyster mushrooms and shiitake mushrooms and 11 kinds of amino acid analyzers There are some reports, such as the determination of 17 free and all amino acids contained in edible mushrooms, the determination of amino acids by TLC and amino acid analyzers, and the author's scent components, organic acids and fatty acid composition. There is a lack of new application products using mushrooms.

Korean Patent Registration Publication No. 10-1479209 (Announcement date 2015.01.09) Korean Patent Registration Publication No. 10-08437886 (Announcement date 2008.07.03)

The present invention is to provide a new product using shiitake mushrooms, shiitake mushroom processed products capable of maximizing the beneficial amino acid content contained in the shiitake mushrooms. That is, the present invention is to provide a method for manufacturing a shiitake mushroom saccharified concentrate, a shiitake mushroom processed product, and a shiitake mushroom saccharified concentrate prepared by the method and a shiitake mushroom saccharified concentrate powder.

In order to solve the above-mentioned problems, the method for preparing saccharified mushroom saccharification concentrate of the present invention is to saccharify the concentrate concentrated by extracting the extract of shiitake mushroom dry powder with saccharification enzyme, and then re-concentrate the saccharide to produce saccharified mushroom saccharification concentrate. The purpose is to provide a method.

In addition, another object of the present invention is to provide a method of manufacturing a shiitake mushroom saccharified concentrated powder further performing a process of lyophilizing and pulverizing the shiitake mushroom saccharified concentrate.

In addition, the present invention has an object to provide a shiitake mushroom saccharification concentrate prepared by the above-mentioned shiitake mushroom saccharification concentrate production method.

In addition, the present invention has an object to provide a shiitake mushroom saccharified concentrated powder produced by the method for producing saccharified mushroom concentrated powder.

In addition, the present invention has an object to provide a health functional food comprising the shiitake mushroom saccharification concentrate and / or shiitake mushroom saccharification concentrate powder.

The shiitake mushroom saccharification concentrate and shiitake mushroom saccharification concentrate powder of the present invention have a very high amino acid content, particularly contain a high concentration of essential amino acids, and have excellent anti-inflammatory effects, and can also be applied to various health functional foods.

1 is a schematic process diagram of a method for producing a saccharified mushroom concentrate of the present invention.
Figure 2 is a schematic process diagram for a method for producing a saccharified concentrate using the ethanol extract of shiitake mushrooms of the present invention.
3 is a result of measuring the cytotoxicity of the saccharified concentrate using Raw water extract of Example 1 and ethanol extract of Example 2 on Raw 264.7 cells.
FIG. 4 is a result of measuring anti-inflammatory properties of the saccharified concentrate using Raw water extract of Example 1 and ethanol extract of Example 2 on Raw 264.7 cells.
5 is a result of measuring the cytotoxicity of Raw 264.7 cells for each of the saccharified concentrated powder Examples 3 to 7 prepared by varying the amount of glycosylation enzyme (50 to 500 g) when producing the saccharified concentrated powder using ethanol extract.
Figure 6 is a result of measuring the anti-inflammatory for Raw 264.7 cells for each of the saccharified concentrated powder Examples 3 to 7 prepared by varying the amount of glycosylation enzyme (50 ~ 500g) when producing saccharified concentrated powder using ethanol extract.

Hereinafter, the present invention will be described in more detail.

The saccharified mushroom saccharification concentrate of the present invention is prepared by saccharifying the concentrate concentrated with the extract of the dried shiitake mushroom powder with saccharifying enzyme, as shown in the schematic process diagram in FIG. Can.

The extract may be hot water extract or ethanol extract.

First, the method for preparing the saccharification concentrate using shiitake mushrooms is as follows.

Preparation method of saccharified concentrate using hot water extract comprises the first step of preparing shiitake mushroom dry powder; A second step of preparing a hot water extract by performing a hot water extraction process on shiitake dried powder; Step 3 to prepare the first concentrate by concentrating the hot water extract under reduced pressure; A fourth step of producing a saccharide by mixing and saccharifying the first concentrate and saccharifying enzyme; And 5 steps of preparing the second concentrate by concentrating the saccharide under reduced pressure.

The dried shiitake mushroom powder is obtained by crushing the dried shiitake mushroom powder after drying the shiitake mushroom. For example, the dried shiitake mushroom powder may be filtered by 40 to 60 mesh.

The hot water extraction process is 1,500 to 2,500 parts by weight of water relative to 100 parts by weight of dried powder of shiitake mushrooms, preferably 1,800 to 2,200 parts by weight of water, more preferably 1,900 to 2,100 parts by weight, and then a temperature of 75 to 90°C. In, preferably, a hot water extraction process may be performed at a temperature of 75 to 85° C. for 10 to 16 hours, preferably for 10 to 14 hours, and more preferably for 11 to 13 hours.

At this time, if the amount of water used is less than 1,500 parts by weight, the shiitake powder may be absorbed and there may be difficulty in filtration. If the amount of water used exceeds 2,500 parts by weight, excessive time and cost may occur in the concentration process due to excessive use. In addition, if the temperature is less than 75°C during hot water extraction, there may be a problem that the amino acid content in the prepared concentrate is low, and when the temperature exceeds 90°C during hot water extraction, there may be a problem that the flavor of shiitake mushroom is lowered. It is preferable to perform a hot water extraction process in the temperature range.

The reduced pressure concentration of the above three steps can be performed by a general method used in the art. The reduced pressure concentration is preferably performed so that the concentration of the first concentrate is 8 to 20 brix, preferably 8 to 15 brix, and more preferably 9 to 13 brix, wherein the concentration of the first concentrate is less than 8 brix. The back side may have a problem of deterioration in the storage process before concentration, and if it exceeds 20 brix, there may be a problem that the temperature control is difficult in the concentration process.

The four-step glycosylation enzyme may be one or more selected from malt, aspergillus oryzae, and black broth (Aspergillus niger), and preferably malt. And, the saccharification is prepared by using 5 to 40 parts by weight of saccharifying enzyme, preferably 10 to 30 parts by weight of saccharifying enzyme, and more preferably 15 to 25 parts by weight based on 100 parts by weight of the first concentrate. It is possible to maximize the essential amino acid content in the saccharified concentrate.

And, the saccharification is a mixture of a first concentrate and a glycosylating enzyme at 55 to 65° C. for 20 to 30 hours, preferably at 57 to 63° C. for 20 to 30 hours, more preferably 58 to 62° C. It is good to obtain a saccharide by fermentation under 20 to 28 hours, and at this time, if the saccharification temperature is less than 55°C or exceeds 65°C, there may be a problem that the essential amino acid content in the saccharide decreases.

Then, the saccharification of shiitake mushrooms using hot water extract is obtained by concentrating the saccharide of step 4 again under reduced pressure to obtain a second concentrate having a concentration of 35 to 50 brix, preferably 35 to 45 brix, and more preferably 37 to 43 brix. A concentrate can be obtained. At this time, the reduced pressure concentration can be performed by a general method used in the art, and in one preferred embodiment, it can be performed by concentrated under reduced pressure under 900hpa ~ 950hpa pressure.

Next, a method of preparing a shiitake mushroom saccharification concentrate using ethanol extract will be described.

The step of preparing a dried powder of shiitake mushrooms is a step of saccharifying mushrooms using ethanol extract; After the ethanol extraction process of the dried shiitake mushroom powder, a second step of removing the solvent to prepare a first ethanol extract; A third step of preparing the first concentrate by concentrating the first ethanol extract under reduced pressure; 4 steps of drying the first concentrate; A fifth step of producing saccharification by mixing and saccharifying the dried first concentrate and saccharifying enzyme; And 6 steps of preparing the second concentrate by concentrating the saccharide under reduced pressure.

The dried shiitake mushroom powder is obtained by crushing the dried shiitake mushroom powder after drying the shiitake mushroom. For example, the dried shiitake mushroom powder may be filtered by 40 to 60 mesh.

In the ethanol extraction process, 1,200 to 2,200 parts by weight of an ethanol aqueous solution, preferably 1,400 to 2,200 parts by weight of an ethanol aqueous solution, more preferably 1,600 to 2,100 parts by weight, and then reflux extraction with respect to 100 parts by weight of dried powder of shiitake mushroom Ethanol extraction process can be performed.

At this time, if the amount of the aqueous ethanol solution is less than 1,200 parts by weight, there may be a problem of gelling, and if the amount of the aqueous solution of the ethanol exceeds 2,200 parts by weight, there is a problem of residual solvent due to excessive use. In addition, the ethanol aqueous solution is preferably an ethanol aqueous solution having a concentration of 70 to 95% by volume, preferably an ethanol aqueous solution having a concentration of 75 to 95% by volume. At this time, if the ethanol aqueous solution concentration is less than 70% by volume, the total amino acid content in the shiitake mushroom saccharification concentrate may be low, and if it exceeds the 95% by volume concentration, there may be a problem that acetaldehyde is generated.

Then, an ethanol aqueous solution as a solvent is removed from the ethanol extract obtained through the ethanol extraction process to prepare a first ethanol extract.

The reduced pressure concentration of the three stages can be performed by a general method used in the art, and the reduced pressure concentration of the first concentrate is 8 to 20 brix, preferably 8 to 15 brix, more preferably 9 to It is good to perform it to be about 13 brix. At this time, if the concentration of the first concentrate is less than 8 brix, there may be a problem of deterioration during storage, and if it exceeds 20 brix, there may be a problem that caramelization occurs. Then, the obtained first concentrate is dried.

The five-step glycation enzyme may use one or more selected from malt, yellow and black soup, and preferably malt. And, the saccharification is prepared by using 5 to 40 parts by weight of glycation enzyme, preferably 10 to 30 parts by weight of glycation enzyme, and more preferably 15 to 25 parts by weight based on 100 parts by weight of the first concentrate. The essential amino acid content in the mushroom saccharification concentrate can be maximized.

And, the saccharification in step 5 is a mixture of the dried first concentrate and saccharifying enzyme for 20 to 30 hours under 55 to 65°C, preferably for 20 to 30 hours under 57 to 63°C, more preferably It is good to obtain a saccharide by fermenting for 20 to 28 hours under 58 to 62°C. At this time, if the saccharification temperature is less than 55°C or exceeds 65°C, there may be a problem that the essential amino acid content in the saccharide decreases.

And, in step 6, the saccharide of step 5 is concentrated again under reduced pressure to obtain a second concentrate having a concentration of 35 to 50 brix, preferably 35 to 45 brix, and more preferably 37 to 43 brix. Shiitake mushroom saccharification concentrate can be obtained. At this time, the reduced pressure concentration may be performed by a general method used in the art, and in one preferred embodiment, it may be performed by concentrated under reduced pressure under 900 hpa to 950 hpa pressure.

According to the present invention, the process of lyophilizing and pulverizing the shiitake mushroom saccharified concentrate prepared above may further be performed to prepare a saccharified mushroom saccharified concentrated powder. When explaining the method for preparing the saccharified mushroom concentrated powder using ethanol extract, as shown in the schematic process diagram in FIG. 2, the second concentrate obtained in step 6 was subjected to an ethanol extraction process to obtain ethanol extract again. 7 steps to do; And 8 steps of lyophilizing and re-pulverizing the obtained ethanol extract to powder it.

At this time, in the ethanol extract of step 7, the second concentrate is mixed with an ethanol aqueous solution having a concentration of 70 to 95% by volume, preferably an ethanol aqueous solution having a concentration of 75 to 95% by volume, and then refluxed to obtain the ethanol extract again. can do.

The shiitake mushroom saccharification concentrate and/or shiitake mushroom saccharification concentrate of the present invention prepared as described above contains 15,500 to 20,000 mg% of total constituent amino acids, more preferably 16,500 to 20,000 mg%, and more preferably 17,000 to 20,000 mg% can do.

And, in the shiitake mushroom saccharification concentrate and/or shiitake mushroom saccharification concentrate of the present invention, the essential constituent amino acid content with respect to the total constituent amino acid content may satisfy Equation 1 below.

[Equation 1]

40% ≤ (essential constituent amino acid content/total constituent amino acid content)×100% ≤ 47%, preferably 41.0% ≤ (essential constituent amino acid content/total constituent amino acid content)×100% ≤ 44.5%, more preferably 42.0 % ≤ (essential constituent amino acid content/total constituent amino acid content)×100% ≤ 44.5%

And, the shiitake mushroom saccharification concentrate and/or shiitake mushroom saccharification concentrate powder of the present invention contains 1,600 to 3,000 mg% of total free amino acids, more preferably 1,800 to 2,800 mg%, and more preferably 2,000 to 2,750 mg%. Can.

In addition, the shiitake mushroom saccharification concentrate and/or shiitake mushroom saccharification concentrate powder of the present invention may satisfy the following equation 2 in terms of the essential free amino acid content relative to the total free amino acid content.

[Equation 2]

40% ≤ (essential free amino acid content/total free amino acid content)×100% ≤ 62%, preferably 45.0% ≤ (essential free amino acid content/total free amino acid content)×100% ≤ 60.0%, more preferably 50.0 % ≤ (essential free amino acid content/total free amino acid content)×100% ≤ 58.0%

The total constituent amino acids and total free amino acids are aspartic acid, serine, glutamic acid, glycine, histidine, arginine, threonine, alanine ), proline, tyrosine, valine, methionine, lysine, isoleucine, leucine and phenylalanine.

The essential constituent amino acids and essential free amino acids may include threonine, valine, methionine, isoleucine, leucine, histidine and lysine.

As mentioned above, the shiitake mushroom saccharification concentrate and/or shiitake mushroom saccharification concentrate of the present invention containing various amino acids in high content also has excellent anti-inflammatory effects, and can be applied as ingredients such as health functional foods to produce various condensed products. It might be.

Hereinafter, the present invention will be described in more detail through examples, but the following examples are not intended to limit the scope of the present invention, which should be interpreted to help understand the present invention.

[ Example ]

Example One : Hydrothermal Shiitake mushrooms through extract Saccharified Produce

1 kg of dried shiitake mushrooms were crushed to prepare a powder that passed through 50 mesh as a sample.

After mixing 1 kg of shiitake mushroom powder and 20 kg of water, reflux extraction was performed at 80° C. for 12 hours to perform hot water extraction to obtain a hot water extract.

Next, the hot water extract was concentrated under reduced pressure from 950 hpa to 900 hpa to obtain a first concentration of 10 brix.

Next, a mixture of the first concentrate and 200 g of malt, a glycosylating enzyme, was fermented at 60° C. for 24 hours to prepare a saccharide.

Next, the saccharide was concentrated under reduced pressure at 950 hpa to 900 hpa to prepare a saccharified mushroom saccharified concentrate, which is a second concentrate having a concentration of 40 brix.

Example 2: Shiitake mushroom through ethanol extract Saccharified Produce

1 kg of dried shiitake mushrooms were crushed to prepare a powder that passed through 50 mesh as a sample.

After mixing 1 kg of shiitake mushroom powder with 20 kg of an ethanol aqueous solution having a concentration of 85% by volume, ethanol extraction was performed by reflux extraction at 30° C. for 12 hours to obtain ethanol extract.

Next, an aqueous ethanol solution was removed from the ethanol extract to prepare an ethanol extract.

Next, the ethanol extract was concentrated under reduced pressure to 900 hpa to obtain a first concentration of 10 brix.

Next, the first concentrate was freeze-dried to obtain a dried first concentrate.

Next, with respect to 100 parts by weight of the dried first concentrate, a mixture of 20 parts by weight of malt that is a glycosylation enzyme was fermented at 60° C. for 24 hours to prepare a saccharide.

Next, the saccharide was concentrated under reduced pressure to 900 hpa to prepare a saccharified mushroom saccharification concentrate, which is a second concentrate having a concentration of 40 brix.

Experimental Example 1: Shiitake mushroom Saccharified Measurement of cytotoxic and anti-inflammatory effects

The cytotoxicity test and anti-inflammatory effect experiment of the shiitake mushroom saccharification solution prepared in Examples 1 and 2 in Raw 264.7 cells were performed.

(1) The cytotoxicity test was performed by incubating Raw 264.7 cells in a 96 well plate at a concentration of 1Х10 ⁵ cells/mL. Then, each of the saccharification solutions of Examples 1 and 2, which are samples, was treated with Raw 264.7 cell culture medium at a concentration of 10, 50, 100, 500 µg/mL, and after 24 hours, the medium was removed and fresh medium was removed. After adding 10 uL MTS to the reaction for 4 hours, absorbance was measured at 540 nm. The cell viability of each sample was calculated relative to the sample-untreated group as 100%, and the results are shown in FIG. 3.

(2) For the anti-inflammatory effect confirmation experiment, Raw 264.7 cells were dispensed into 96 well plates at a concentration of 1Х10 ⁵ cells/mL and cultured for 24 hours. Next, it was treated with saccharification solutions of Example 1 and Example 2 and LPS (Lipopolysaccharide). At this time, LPS was treated at a concentration of 1 μg/mL, respectively, and each sample, each of the saccharification solutions of Examples 1 and 2, was diluted to a concentration of 10, 50, 100, 500 μg/mL and added to the cells.

After 24 hours, the cell culture solution was collected, and the ability to inhibit NO production was measured using a NO assay using the Griess reagent system, and the results are shown in FIG. 4.

Referring to Figure 3, when applied at a concentration of 500μg / mL showed a cytotoxic result. On the other hand, there was no cytotoxicity at 50-100 μg/mL.

Referring to Figure 4, as the saccharification solution concentration increased, it was found that the NO production inhibitory ability significantly increased. In addition, the saccharification solution by the ethanol extract of Example 2 showed better results in inhibiting NO production than the saccharification solution by the hot water extract of Example 1.

Through the above experiment, the saccharification solution using the hot water extract or the ethanol extract of the present invention has an excellent NO generation inhibitory ability without cytotoxicity when applied at a concentration of less than 500 μg/mL, preferably at a concentration of 50 to 400 μg/mL. I could confirm.

Example 3-1: Shiitake mushrooms through ethanol extract Saccharified Produce

1 kg of dried shiitake mushrooms were crushed to prepare a powder that passed through 50 mesh as a sample.

After mixing 1 kg of shiitake mushroom powder and 20 kg of an ethanol aqueous solution having a concentration of 85% by volume, reflux extraction was performed at 30° C. for 12 hours to perform ethanol extraction to obtain an ethanol extract.

Next, a first ethanol extract was prepared by removing the aqueous ethanol solution as a solvent from the ethanol extract.

Next, the first ethanol extract was concentrated under reduced pressure to 950 hpa to obtain a first concentration of 10 brix.

Next, the first concentrate was freeze-dried to obtain a dried first concentrate.

Next, with respect to 100 parts by weight of the dried first concentrate, a mixture of 20 parts by weight of malt that is a glycosylation enzyme was fermented at 60° C. for 24 hours to prepare a saccharide.

Next, the saccharide was concentrated under reduced pressure to 950 hpa to prepare a second concentrate having a concentration of 40 brix.

Next, the second concentrate was mixed with an aqueous ethanol solution having a concentration of 90% by volume, and reflux was extracted to prepare a second ethanol extract.

Next, the second ethanol extract was freeze-dried at -40°C, and then pulverized to prepare saccharified mushroom saccharified concentrated powder.

Example 3-2 ~ Example 3-5

In the same manner as in Example 3-1, shiitake mushroom glycated concentrated powder was prepared, but the amount of malt used as glycation enzyme at the time of saccharification was varied as shown in Table 1 below, to prepare shiitake mushroom glycated concentrated powder, respectively, Examples 3-2 to 3 -5 was performed respectively.

division First concentrate (parts by weight) Glycosylase
(Malt, parts by weight) Example 3-1 (A) 100 5 Example 3-2 (B) 100 10 Example 3-3 (C) 100 20 Example 3-4 (D) 100 30 Example 3-5 (E) 100 50

Experimental Example 2: Shiitake mushroom Saccharified Measurement of cytotoxic and anti-inflammatory effects

Cytotoxicity test and anti-inflammatory effect test on raw 264.7 cells of shiitake mushroom saccharide powder prepared in Examples 3-1 to 3-5 were performed.

Cytotoxicity test was used in the experiment by culturing Raw 264.7 cells in a 96 well plate at a concentration of 1×10 ⁵ cells/mL. Then, each of the saccharified concentrated powders of Examples 3-1 to 3-5, which is a sample in a raw 264.7 cell culture medium, was treated at a concentration of 10, 50, 100, 500 μg/mL, and after 24 hours, the medium was removed. Then, after adding 10 uL MTS to the new medium and reacting for 4 hours, absorbance was measured at 540 nm. The cell viability of each sample was calculated relative to the sample-untreated group as 100%, and the results are shown in FIG. 5 and Table 2 below.

Cell viability
(%) Addition amount of saccharified concentrated powder 10
μg/mL 50
μg/mL 100
μg/mL 500
μg/mL Example 3-1 (A) 98.4% 96.4 94.9 92.8 Example 3-2 (B) 93.5 90.2 89.1 82.4 Example 3-3 (C) 98.5 95.2 93.3 90.9 Example 3-4 (D) 95.8 91.0 88.6 86.3 Example 3-5(E) 88.3 83.2 75.3 66.3

In the case of Examples 3-5, the cell viability was 75.3% at a concentration of 100 µg/mL and 66.3% at a concentration of 500 µg/mL, indicating toxicity. Unlike Examples 3-1 to 3-4, overall low Showed cell viability. In contrast, in the case of Examples 3-1 to 3-4, the cell viability was 86% or more, preferably 90% or more, and in particular, Examples 3-1 and 3-3 were added to the saccharified concentrated powder range. All showed high cell viability of over 90%.

(2) For the anti-inflammatory effect confirmation experiment, Raw 264.7 cells were dispensed into 96 well plates at a concentration of 1Х10 ⁵ cells/mL and cultured for 24 hours. Next, it was treated with the glycated concentrated powder of Example 3-1 to 3-5 and LPS (Lipopolysaccharide). At this time, LPS was treated at a concentration of 1 μg/mL, respectively, and each of the glycated concentrated powders of Examples 3-1 to 3-52, which were samples, was diluted to a concentration of 10, 50, 100, 500 μg/mL, and applied to the cells. Was added.

After 24 hours, the cell culture solution was recovered, and the NO generation inhibitory ability was measured using a NO assay using the Griess reagent system, and the results are shown in FIG. 6.

Referring to FIG. 6, when Example 3-3(C) was treated at a concentration of 500 μg/mL, NO generation rate of 76.9% was shown, and in Example 3-5(E), 71.4 at a concentration of 500 μg/mL. % NO production rate, but it seems to be due to the decrease in cell viability due to toxicity. Example 3-1(A), Example 3-2(B), and Example 3-4(D) were treated with a concentration of 500 μg/mL, respectively, resulting in NO production rates of 94.1%, 83.1%, and 80.2%. Showed.

Experimental Example 3: Shiitake mushroom Saccharified concentrated powder Composition amino acid and free amino acid measurement

The constituent amino acids and free amino acids of each of the shiitake mushroom saccharified concentrated powders of Examples 3-1 to 3-5 were measured in the following manner, and the results are shown in Tables 4 and 5, respectively.

(1) Constituent amino acid

Constitutive amino acid analysis, 0.5 g of sample crushed sample is put into a test tube, 10 N of 6 N-HCl is added, the end of the test tube is melted to form an ampoule, sealed, hydrolyzed at 110°C for 24 hours, and then the ampoule is broken and filtered with filter paper. While filtrating, it was dissolved in 50 mL of methanol, concentrated under reduced pressure, and dissolved in 5 mL of 20 mM HCl. Next, a certain amount of the filtrate obtained by filtration with a 0.45 μm membrane filter was taken and derivatized using AccQ-Tag reagent, and then analyzed by HPLC. Table 3 shows the analysis conditions.

(2) free amino acids

For free amino acid analysis, 25 mg of sulfosalicylic acid was added to 10 mL of the filtrate obtained in the same way as the determination of free sugar, and left to stand at 4°C for 4 hours, followed by centrifugation (50000 rpm, 30 minutes) to remove proteins and the like. , The supernatant was filtered with a 0.45 μm membrane filter, and a certain amount of the filtrate was derivatized using AccQ-Tag reagent and analyzed by HPLC. Table 3 shows the analysis conditions.

Item Analysis conditions device Agilent Technologies 1200 Series Detector Agilent Technologies 1200 Series FLD column AccQ-Tag?? (Waters Co., 150 mm L×3.9 mm I.D.) Column temperature 37℃ Buffer solution
(Buffer solution)

A: AccQ-Tag Eluent A (acetate-phosphate buffer)
B: AccQ-Tag Eluent B (100% acetonitrile)
C: DW (Distilled water)

flux 1.0 mL/min Injection volume 5 μL

Configuration
amino acid
(mg%) Example
3-1 Example
3-2 Example
3-3 Example
3-4 Example
3-5 Aspartic acid 580.51±90.29 ⁴⁾ 670.70±98.64 892.32±148.92 520.07±13.62 340.45±27.42 Serine 1,129.91±171.34 1,056.33±54.64 1,248.67±98.96 1,063.44±13.67 882.62±44.40 Glutamic acid 2,494.07±385.59 2,536.40±252.17 2,819.69±344.82 2,222.43±7.12 1,479.30±80.82 Glycine 808.77±117.99 792.74±53.02 990.09±97.98 770.22±8.04 586.75±37.67 Histidine 873.64±128.45 960.89±58.55 990.02±79.49 850.22±29.26 855.83±49.90 Arginine 3,719.45±553.89 3,778.64±166.94 4,122.51±293.38 3,617.84±91.31 3,288.80±108.35 Threonine 1,101.95±168.61 1,014.08±72.09 1,298.33±122.92 1,012.61±17.23 715.53±64.71 Alanine 166.81±9.47 143.00±7.09 169.93±10.08 156.32±0.91 124.04±3.55 Proline 575.87±80.45 536.50±22.78 642.84±45.89 577.77±5.14 448.82±31.85 Tyrosine 408.41±54.14 484.48±33.10 504.48±40.50 389.57±19.04 344.39±2.08 Valine 1,352.82±203.42 1,081.68±16.25 1,211.05±74.61 1,213.95±8.07 1,173.39±40.77 Methionine 232.30±33.64 251.79±20.81 277.92±26.82 222.34±18.75 212.72±14.05 Lysine 686.51±114.36 681.97±103.77 844.24±132.74 522.47±57.86 272.57±39.42 Isoleucine 999.83±149.61 896.51±21.33 1,089.08±56.25 951.10±17.20 726.71±55.69 Leucine 1,049.79±158.54 1,069.11±96.19 1,349.35±145.10 998.12±15.60 785.04±53.51 Phenylalanine 1,050.53±143.42 1,197.54±70.60 1,248.51±103.10 1,057.02±67.88 1,131.03±20.56 TAA ^{1 )} 17,231.14±563.22 17,152.38±573.99 19,699.01±821.54 16,145.49±120.30 13,367.98±674.75 EAA ^{2 )} 7,347.35±502.05 7,153.57±459.60 8,308.48±541.01 6,827.83±31.13 5,872.80±338.60 EAA/TAA(%) ^{3 )} 42.64±0.26 41.77±0.12 42.29±0.15 42.29±0.12 43.90±0.32 ¹⁾ TAA: Total amino acid
²⁾ EAA: Essential amino acid=Tyrosine + Valine + Methionine + Isoleucine + Leucine + Histidine + Lysine
³⁾ EAA/TAA (%) is measured according to Equation 1 below.
[Equation 1]
(Essential constituent amino acid content/total constituent amino acid content)×100%
⁴⁾ Average value (±SD)

Amino acids are important nutrients that make up living organisms and must be consumed externally to maintain them. The major amino acids of the 16 amino acids were arginine, glutamic acid, and serine, and the total amino acid content of Example 3-3 was the highest at 19,699.01 mg%. 17,231.14 mg% in Example 3-1 and 17,152.38 mg% in Example 3-2 were highest, and Example 3-5 was the lowest at 13,367.98 mg%.

The main amino acids were arginine, a basic amino acid, and glutamic acid, a type of acidic amino acid. Arginine content is 4,122.51 mg% in Example 3-3, 3,778.64 mg% in Example 3-2, 3,719.45 mg% in Example 3-1, 3,617.84 mg% in Example 3-4, Example 3- 5 to 3,288.80 mg%.

And, the glutamic acid content is 2,819.69 mg% in Example 3-3, 2,536.40 mg% in Example 3-2, 2,494.09 mg% in Example 3-1, 2,222.43 mg% in Example 3-4, and implementation In Example 3-5, the order was 1,479.30 mg%.

The composition amino acid content of shiitake mushroom glycated concentrated powder according to the amount of glycation enzyme was highest in Example 3-3 using 20 parts by weight of glycase, and the content of arginine in major amino acids related to immunity was also highest in Example 3-3. .

And, EAA/TAA (%) is 40% or more, preferably 41 ~ 44.5%, more preferably 42 ~ 44.5%, the ratio of essential amino acid content was very high.

Free amino acids
(mg%) Example
3-1 Example
3-2 Example
3-3 Example
3-4 Example
3-5 Aspartic acid 53.00±2.37 ⁴⁾ 34.98±4.00 35.03±2.80 47.55±5.30 41.26±2.36 Serine 64.60±0.44 125.99±5.52 127.43±3.82 75.72±4.24 17.39±1.26 Glutamic acid 526.96±21.47 352.25±43.81 505.56±35.10 389.53±58.58 149.36±10.02 Glycine 45.39±0.46 54.97±1.92 50.96±0.92 27.88±1.42 11.74±0.74 Histidine 452.01±10.08 679.99±33.79 931.79±34.13 723.51±40.10 19.05±1.17 Arginine 67.67±2.94 207.20±4.33 167.12±7.25 127.92±4.06 48.32±2.47 Threonine 64.69±0.31 121.25±5.42 113.44±3.94 71.56±3.25 69.47±4.21 Alanine 136.98±3.33 130.30±13.44 136.19±8.01 96.63±9.72 117.27±8.64 Proline 21.06±1.51 28.84±1.49 18.62±0.08 26.95±2.09 31.17±2.75 Tyrosine 25.98±0.82 42.52±8.70 51.57±0.89 40.56±0.35 8.41±0.64 Valine 41.38±1.60 84.06±2.52 68.14±3.96 44.52±2.08 43.27±1.48 Methionine 8.08±1.06 17.90±1.17 15.04±1.19 10.98±0.80 11.71±1.27 Lysine 61.85±2.29 125.43±11.66 130.61±11.33 88.46±9.78 48.07±3.21 Isoleucine 28.18±1.75 53.30±0.50 41.80±3.02 27.94±0.24 16.08±1.24 Leucine 33.98±2.92 83.39±1.02 68.49±5.20 41.56±0.46 36.27±3.27 Phenylalanine 41.00±5.30 100.80±4.05 79.87±5.30 54.00±3.80 21.64±2.16 TAA ^{1 )} 1,672.82±1.89 2,243.16±129.86 2,541.67±126.92 1,895.29±136.16 690.48±42.19 EAA ^{2 )} 731.18±24.69 1,266.12±46.65 1,449.19±68.05 1,062.54±50.37 265.56±19.64 EAA/TAA(%) ^{3 )} 43.71±1.53 56.48±1.19 57.02±0.17 56.11±1.37 39.08±0.11 ¹⁾ TAA: Total amino acid
²⁾ EAA: essential amino acid=Tyrosine + Valine + Methionine + Isoleucine + Leucine + Histidine + Lysine
³⁾ EAA/TAA (%) is measured according to Equation 2 below.
[Equation 2]
(Essential free amino acid content/total free amino acid content)×100%
⁴⁾ Average value (±SD)

Looking at Table 5, a total of 16 free amino acids were detected in all the shiitake mushroom saccharified concentrated powders of Examples 3-1 to 3-5, and the total free amino acids were the highest 2,541.67 mg% in Example 3-3. 2,243.16 mg% in Example 3-2, 1,895.29 mg% in Example 3-4, and 1,672.82 mg% in Example 3-1. In addition, histidine and glutamic acid were the highest as the main amino acids.

In addition, in Example 3-3 to which 20 parts by weight of glycation enzyme (malt) was applied, the essential amino acid content was highest.

And, EAA/TAA (%) is 40% or more, preferably 45 to 60%, more preferably 50 to 58%, the essential free amino acid content ratio was very high.

Through the above Examples and Experimental Examples, it was confirmed that the shiitake mushroom saccharification concentrate and shiitake mushroom saccharification concentrated powder prepared by the method of the present invention have excellent anti-inflammatory properties and have a very high essential amino acid content.

Claims (14)

A method for producing saccharose mushroom saccharification concentrate, characterized in that after the saccharification of the concentrated concentrate of dried shiitake mushroom extract by saccharification, the saccharide is reconcentrated.
According to claim 1, If the extract is a hot water extract,
Preparing shiitake mushroom dry powder in 1 step;
A second step of preparing a hot water extract by performing a hot water extraction process on shiitake dried powder;
Step 3 to prepare the first concentrate by concentrating the hot water extract under reduced pressure;
A fourth step of producing a saccharide by mixing and saccharifying the first concentrate and saccharifying enzyme; And
A fifth step of preparing the second concentrate by concentrating the saccharide under reduced pressure;
A method of manufacturing shiitake mushroom saccharification concentrate comprising a.
According to claim 1, If the extract is an ethanol extract,
Preparing shiitake mushroom dry powder in 1 step;
After the ethanol extraction process of the dried shiitake mushroom powder, a second step of removing the solvent to prepare a first ethanol extract;
A third step of preparing the first concentrate by concentrating the first ethanol extract under reduced pressure;
4 steps of drying the first concentrate;
A fifth step of producing saccharification by mixing and saccharifying the dried first concentrate and saccharifying enzyme; And
Step 6 to produce a second concentrate by concentrating the sugar under reduced pressure;
A method of manufacturing shiitake mushroom saccharification concentrate comprising a.
The method of claim 2 or 3, wherein the first concentrate has a concentration of 8 to 20 brix, and the second concentrate has a concentration of 35 to 50 brix.
The method of claim 2 or 3, wherein the saccharification is performed by mixing 5 to 40 parts by weight of saccharifying enzyme with respect to 100 parts by weight of the first concentrate.
The method of claim 5, wherein the saccharifying enzyme comprises at least one selected from malt, malt, Aspergillus oryzae and Aspergillus niger, the saccharification is performed for 20 to 30 hours at 55 to 65°C. A method of manufacturing shiitake mushroom saccharification concentrate, characterized in that.
The method of claim 2, wherein the hot water extraction process is characterized in that after mixing 1,500 to 2,500 parts by weight of water with respect to 100 parts by weight of shiitake mushroom dry powder, the hot water extraction process is performed at a temperature of 75 to 90°C for 10 to 16 hours. Preparation method of shiitake mushroom saccharification concentrate.
The method of claim 3, wherein the ethanol extraction process comprises mixing 1,200 to 2,200 parts by weight of an ethanol aqueous solution having a concentration of 90% by volume with respect to 100 parts by weight of dried powder of shiitake mushrooms, and then extracting ethanol at a temperature of 25 to 30°C for 8 to 12 hours. A method of producing shiitake mushroom saccharification concentrate, characterized by performing a process.
A method for producing a saccharified mushroom saccharified concentrated powder according to claim 2 or 3, wherein the second concentrate is freeze-dried and pulverized.
A method for producing saccharified mushroom saccharified concentrated powder, characterized in that the second concentrate of claim 3 is subjected to an ethanol extraction process to obtain the ethanol extract again, and then freeze-dried and ground to powder.
Shiitake mushroom saccharification concentrate, characterized in that produced by the method of claim 2 or 3.
Claim 9, characterized in that produced by the method of shiitake mushroom saccharified concentrated powder.
The method of claim 12, wherein the essential constituent amino acid content relative to the total constituent amino acid content satisfies Equation 1 below,
The essential constituent amino acids include threonine, valine, methionine, isoleucine, leucine, histidine and lysine,
The total constituent amino acids are aspartic acid, serine, glutamic acid, glycine, histidine, arginine, threonine, alanine, proline ( shiitake mushroom saccharification concentrate powder comprising proline, tyrosine, valine, methionine, lysine, isoleucine, leucine and phenylalanine;
[Equation 1]
40% ≤ (essential constituent amino acid content/total constituent amino acid content)×100% ≤47%
The method of claim 12, wherein the essential free amino acid content relative to the total free amino acid content satisfies Equation 2,
The essential free amino acids include threonine, valine, methionine, isoleucine, leucine, histidine and lysine,
The total free amino acids are aspartic acid, serine, glutamic acid, glycine, histidine, arginine, threonine, alanine, proline ( shiitake mushroom saccharification concentrate powder comprising proline, tyrosine, valine, methionine, lysine, isoleucine, leucine and phenylalanine;
[Equation 2]
40% ≤ (essential free amino acid content/total free amino acid content)×100% ≤ 62%

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