TWI750084B - Method of extracting active ingredients in mushrooms - Google Patents

Abstract

A method of extracting active ingredients in mushrooms includes following steps: crushing a mushroom to obtain a crude mushroom crush; mixing the crude mushroom crush with deionized water to obtain a crude crush mixture; adding the crude crush mixture into a subcritical fluid extraction machine and extracting the crude crush mixture to obtain a crude mushroom extract; centrifuging the crude mushroom extract to harvest a first supernatant and a mushroom residue; mixing the mushroom residue with deionized water and a cellulase enzyme to obtain a reactive mixture; adding the reactive mixture into a high-pressure hydrolysis reactor and extracting the reactive mixture to obtain a hydrolysate; centrifuging the hydrolysate to obtain a second supernatant; and combining the first supernatant and the second supernatant, and then concentrating the combined first and second supernatants to obtain a mushroom extraction solution.

Description

Method for extracting active ingredients from mushrooms

The present invention relates to an extraction method of mushrooms, and particularly relates to an extraction method capable of efficiently extracting active ingredients of mushrooms.

Mushrooms contain a variety of special ingredients (such as polymer polysaccharides, triterpenoids, unsaturated fatty acids, trace elements, flavonoids, etc.), which can enhance and activate the immune system, prevent cancer, lower blood sugar, lower blood lipids, antioxidants, Anti-pneumonia, anti-dementia, analgesia, and enhancement of liver function and other benefits.

Although mushrooms have many functional components that are beneficial to the human body, most of them are stored in the cells, and the composition of the cell wall is mainly tough chitin. The structure of chitin is complex, and the human body is difficult to digest, which hinders the release and release of effective components. absorb. The traditional method of extracting mushrooms is to use hot water extraction method, which has the problems of long cooking time, high temperature and low yield. In order to overcome the shortcomings of traditional hot water extraction methods, in recent years, new extraction technologies and methods such as ultrasonic or microwave assisted extraction, supercritical, and enzyme hydrolysis have been gradually derived, but the extraction effects of these methods are also very unsatisfactory, and they will be used. To a large amount of organic solvents such as ethanol.

Therefore, there is still an urgent need to provide a method for extracting active ingredients from mushrooms, which can increase the yield of active ingredients in mushroom extracts and avoid the problems of long boiling time, high temperature, and low yield in traditional extraction methods. , And do not need to use a lot of organic solvents.

The purpose of the present invention is to provide a method for extracting active ingredients from mushrooms. Compared with the prior art, the method for extracting active ingredients from mushrooms of the present invention can achieve the effect of increasing the yield of active ingredients in mushroom extracts, and can avoid traditional extraction methods with long boiling time, high temperature and yield. For low-quality problems, there is no need to use a lot of organic solvents.

In order to achieve the above objective, the present invention provides a method for extracting active ingredients from mushrooms, which includes the following steps: a crushing step, crushing the mushrooms with a coarse crusher to obtain a coarse mushrooms; Step: Mix the coarsely chopped mushrooms and deionized water at a mass-volume ratio of 1:400 to 10:400 to obtain a coarsely chopped mixture; in a subcritical extraction step, add the coarsely chopped mixture to The extraction is carried out in a subcritical extraction tank. The pressure of the subcritical extraction tank is between 1000 and 5000 psi, the extraction temperature is between 50 and 200°C, and the extraction time is between 5 and 40 minutes. Crude mushroom extract: a first centrifugation step, centrifuge the crude mushroom extract at 5000 rpm to 15000 rpm for 5 to 30 minutes, and collect the supernatant separated from the crude mushroom extract to obtain A first supernatant, and collecting the lower sediment separated from the crude mushroom extract to obtain a mushroom residue; a mushroom residue mixing step, mixing the mushroom residue with a deionized water and a Cellulose hydrolase is uniformly mixed to obtain a reaction mixture; in a high-pressure hydrolysis step, the reaction mixture is added to a high-pressure hydrolysis tank for extraction. The pressure of the high-pressure hydrolysis tank is between 500 and 5000 psi, and the extraction temperature is between The extraction time is between 10 and 100°C, and the extraction time is between 10 and 100 minutes. After the extraction is completed, a hydrolysate is obtained; a second centrifugation step, the hydrolysate is centrifuged at 5000 rpm to 15000 rpm for 5 to 30 minutes, Collect the supernatant separated from the hydrolysate to obtain a second supernatant; and a vacuum concentration step in which the first supernatant and the second supernatant are mixed and concentrated under reduced pressure at 37°C to obtain A mushroom extract.

In one embodiment, in the mixing step, the mass-volume ratio of the crude mushrooms to the deionized water is between 3:400 and 9:400.

In one embodiment, in the subcritical extraction step, the pressure of the subcritical extraction tank is between 1500 and 4500 psi, the extraction temperature is between 100 and 150° C., and the extraction time is between 10 and 30 minutes.

In one embodiment, in the high-pressure hydrolysis step, the pressure of the high-pressure hydrolysis tank is between 1000 and 3000 psi, the extraction temperature is between 30 and 60° C., and the extraction time is between 30 and 90 minutes.

In one embodiment, in the first centrifugation step, the crude mushroom extract is centrifuged at a speed of 6000 rpm to 13000 rpm for 10 to 20 minutes.

In one embodiment, in the second centrifugation step, the hydrolysate is centrifuged at a rotation speed of 6000 rpm to 13000 rpm for 10 to 20 minutes.

In one embodiment, in the crushing step, the mushrooms are crushed with a coarse crusher for 10 to 50 minutes.

In one embodiment, in the crushing step, the mushrooms are crushed by a coarse crusher for 20 to 30 minutes.

In one embodiment, the particle size of the coarsely divided mushrooms is between 0.1 and 3 mm.

In one embodiment, the size of the coarsely divided mushrooms is between 0.5 and 2 mm.

In one embodiment, in the mushroom residue mixing step, the mushroom residue and deionized water are mixed in a mass-volume ratio of 1:20 to 1:250, and then cellulose with a concentration of 0.05 to 1% is added. The hydrolase is evenly mixed.

In one embodiment, in the mushroom residue mixing step, the mushroom residue and deionized water are mixed in a mass-volume ratio of 1:50 to 1:200, and then cellulose with a concentration of 0.1 to 0.6% is added. The hydrolase is evenly mixed.

In one embodiment, the mushroom extract is rich in β- glucan and triterpenes.

In one embodiment, the mushroom is Brazilian mushroom, Phellinus igniarius or Tremella.

In summary, the method for extracting active ingredients from mushrooms of the present invention can achieve the effect of increasing the yield of active ingredients in mushroom extracts, and can avoid traditional extraction methods with long boiling time, high temperature, and low yield. And other problems, it is not necessary to use a large amount of organic solvents.

Hereinafter, preferred embodiments and experimental examples of the method for extracting active ingredients from mushrooms according to the present invention will be described with reference to related drawings and tables, in which the same elements will be described with the same reference signs.

The method for extracting active ingredients from mushrooms of the present invention can achieve the effect of increasing the yield of active ingredients in mushroom extracts, and can avoid the problems of long boiling time, high temperature and low yield in traditional extraction methods, not to mention A large amount of organic solvents must be used.

In the present invention, the mushrooms can be Brazilian mushrooms, Phellinus igniarius, Tremella, or other mushrooms commonly known to those skilled in the art.

Brazilian mushroom ( Agaricus blazei Murill , also known as Agaricus subrufescens , or almond mushroom, or Brazil mushroom) is a kind of edible and medicinal mushroom, also known as Brazilian mushroom, Agaricus blazei, sun mushroom, god of life, mushroom of life, Brazil, Peru, with sweet taste and almond aroma. Studies have shown that Brazilian mushrooms contain a variety of special ingredients (such as polymer polysaccharides, triterpenoids, unsaturated fatty acids, trace elements, etc.), which can enhance and activate the immune system, prevent cancer, lower blood sugar, lower blood lipids, and resist oxidation. And to improve liver function and other benefits.

Phellinus linteus ( Phellinus linteus ) is a kind of edible and medicinal mushrooms, also known as mulberry ears and sangchen, which grow on the trunks of Morus plants. Studies have shown that Phellinus igniarius contains a variety of special ingredients (such as polymer polysaccharides, triterpenoids, flavonoids, unsaturated fatty acids, trace elements, etc.), which can enhance and activate the immune system, prevent cancer, prevent cardiovascular disease, and resist Allergies, anti-dementia, soothing sleep, analgesia and improving liver function and other benefits.

Tremella fuciformis ( Tremella fuciformis ) is a kind of edible and medicinal mushrooms, also known as white fungus and white fungus. It has a translucent chrysanthemum shape or chicken crown. Studies have shown that Tremella contains a variety of special ingredients (such as polymer polysaccharides, triterpenes, trehalose, pentose, mannitol, unsaturated fatty acids, trace elements, etc.), and its nutritional value is high, and more It can enhance and activate the immune system, respiratory system, central nervous system, blood circulation system and other benefits.

Please refer to FIG. 1 and FIG. 2. FIG. 1 is a schematic diagram of a preferred embodiment of the method for extracting active ingredients from mushrooms of the present invention. Fig. 2 is a flow chart of a preferred embodiment of the method for extracting active ingredients from mushrooms of the present invention. In this embodiment, the method for extracting active ingredients of mushrooms includes the following steps: a crushing step, crushing the mushrooms with a coarse crusher to obtain a coarse mushrooms (step S01); Step: Mix the coarsely chopped mushrooms with a deionized water in a mass-volume ratio of 1:400 to 10:400 to obtain a coarsely chopped mixture (step S02); a subcritical extraction step, the crude The crushed mixture is added to a subcritical extraction tank for extraction. The pressure of the subcritical extraction tank is between 1000 and 5000 psi, the extraction temperature is between 50 and 200°C, and the extraction time is between 5 and 40 minutes. After the extraction, a crude mushroom extract is obtained (step S03); a first centrifugation step, the crude mushroom extract is centrifuged at 5000 rpm to 15000 rpm for 5 to 30 minutes, and the crude mushroom extract is collected The supernatant liquid separated from the liquid obtains a first supernatant liquid, and the lower sediment separated from the crude mushroom extract is collected to obtain a mushroom residue (step S04); a mushroom residue mixing step is The mushroom residues are uniformly mixed with a deionized water and a cellulose hydrolyzing enzyme to obtain a reaction mixture (step S05); a high-pressure hydrolysis step, the reaction mixture is added to a high-pressure hydrolysis tank for extraction, and high-pressure hydrolysis The pressure of the tank is between 500 and 5000 psi, the extraction temperature is between 10 and 100°C, and the extraction time is between 10 and 100 minutes. After the extraction, a hydrolysate is obtained (step S06); a second centrifugation step Centrifuge the hydrolyzed liquid at a speed of 5000 rpm to 15000 rpm for 5 to 30 minutes, and collect the supernatant separated from the hydrolyzed liquid to obtain a second supernatant (step S07); The first supernatant and the second supernatant are mixed, and concentrated under reduced pressure at 37° C. to obtain a mushroom extract (step S08).

In this embodiment, in the mixing step (step S02), the crude mushrooms and deionized water can be mixed uniformly in a mass-volume ratio of 1:400 to 10:400. Preferably, the mass-volume ratio of coarse mushrooms and deionized water may be between 3:400 and 9:400. Preferably, the mass-volume ratio of coarse mushrooms and deionized water can be 3:400, 4:400, 5:400, 6:400, 7:400, 8:400, 9:400 or any of the foregoing. Any value and range covered between two ratios.

In this embodiment, in the subcritical extraction step (step S03), the pressure of the subcritical extraction tank can be set to be between 1000 and 5000 psi, and the extraction temperature can be set to be between 50 and 200°C. The time can be set between 5 and 40 minutes. Preferably, the pressure of the subcritical extraction tank can be set between 1500 and 4500 psi, the extraction temperature can be set between 100 and 150°C, and the extraction time can be set between 10 and 30 minutes. Preferably, the pressure of the subcritical extraction tank can be set to 1500 psi, 2000 psi, 2500 psi, 3000 psi, 3500 psi, 4000 psi, 4500 psi or any value and range covered by any two of the foregoing values. Preferably, the extraction temperature of the subcritical extraction tank can be set to 100°C, 105°C, 110°C, 115°C, 120°C, 125°C, 130°C, 135°C, 140°C, 145°C, 150°C or any two of the foregoing. Any number and range covered by the number. Preferably, the extraction time of the subcritical extraction tank can be set to 10 minutes, 15 minutes, 20 minutes, 25 minutes, 30 minutes, or any value and range covered by any two of the foregoing values. The setting conditions of the aforementioned subcritical extraction tank are not a limitation of the present invention, as long as the active ingredients can be fully extracted.

In this embodiment, in the high-pressure hydrolysis step (step S06), the pressure of the high-pressure hydrolysis tank can be set between 500 and 5000 psi, the extraction temperature can be set between 10 and 100°C, and the extraction time can be Set between 10 and 100 minutes. Preferably, the pressure of the high-pressure hydrolysis tank can be set between 1000 and 3000 psi, the extraction temperature can be set between 30 and 60°C, and the extraction time can be set between 30 and 90 minutes. Preferably, the pressure of the high-pressure hydrolysis tank can be set to 1000 psi, 1500 psi, 2000 psi, 2500 psi, 3000 psi, or any value and range covered by any two of the foregoing values. Preferably, the extraction temperature of the high-pressure hydrolysis tank can be set to 30°C, 35°C, 40°C, 45°C, 50°C, 55°C, 60°C, or any value and range encompassed between any two of the foregoing values. Preferably, the extraction time of the high-pressure hydrolysis tank can be set to 30 minutes, 35 minutes, 40 minutes, 45 minutes, 50 minutes, 55 minutes, 60 minutes, 65 minutes, 70 minutes, 75 minutes, 80 minutes, 85 minutes, 90 minutes. Minutes or any value and range encompassed between any two of the foregoing values. The setting conditions of the aforementioned high-pressure hydrolysis tank are not a limitation of the present invention, as long as the reaction mixture can be fully hydrolyzed.

In this embodiment, in the first centrifugation step (step S04), the crude mushroom extract can be centrifuged at a rotation speed of 5000 rpm to 15000 rpm for 5 to 30 minutes. Preferably, the crude mushroom extract can be centrifuged at a speed of 6000 rpm to 13000 rpm for 10 to 20 minutes. Preferably, the crude extract of mushrooms can be at 6000 rpm, 6500 rpm, 7000 rpm, 7500 rpm, 8000 rpm, 8500 rpm, 9000 rpm, 9500 rpm, 10000 rpm, 10500 rpm, 11000 rpm, 11500 rpm, 12000 Centrifuge for 10 minutes, 11 minutes, 12 minutes, 13 minutes, 14 minutes, 15 minutes, 16 minutes, 17 minutes at a speed of rpm, 12500 rpm, 13000 rpm or any value and range covered by any two of the foregoing values , 18 minutes, 19 minutes, 20 minutes, or any value and range covered by any two of the foregoing values. The aforementioned setting conditions of centrifugation are not a limitation of the present invention, as long as the supernatant liquid and the lower sediment can be sufficiently separated.

In this embodiment, in the second centrifugation step (step S07), the hydrolyzed solution may be centrifuged at a rotation speed of 5000 rpm to 15000 rpm for 5 to 30 minutes. Preferably, the hydrolysate can be centrifuged at a speed of 6000 rpm to 13000 rpm for 10 to 20 minutes. Preferably, the hydrolyzed liquid can be at 6000 rpm, 6500 rpm, 7000 rpm, 7500 rpm, 8000 rpm, 8500 rpm, 9000 rpm, 9500 rpm, 10000 rpm, 10500 rpm, 11000 rpm, 11500 rpm, 12000 rpm, 12500 rpm Centrifuge for 10 minutes, 11 minutes, 12 minutes, 13 minutes, 14 minutes, 15 minutes, 16 minutes, 17 minutes, 18 minutes, 13000 rpm or any value and range between any two of the foregoing values. 19 minutes, 20 minutes, or any value and range covered by any two of the foregoing values. The aforementioned setting conditions of centrifugation are not a limitation of the present invention, as long as the supernatant liquid and the lower sediment can be sufficiently separated.

In this embodiment, in the crushing step (step S01), the mushrooms can be crushed with a coarse crusher for 10 to 50 minutes. Preferably, the mushrooms can be crushed with a coarse crusher for 20 to 30 minutes. Preferably, the time for the crushing treatment can be 20 minutes, 21 minutes, 22 minutes, 23 minutes, 24 minutes, 25 minutes, 26 minutes, 27 minutes, 28 minutes, 29 minutes, 30 minutes or any two of the foregoing values. Any value and range covered by the space is sufficient so that the size of the coarse mushrooms is less than or equal to 3 mm, so as to facilitate the subsequent subcritical extraction.

In this embodiment, the particle size of the crude mushrooms may be between 0.1 and 3 mm. Preferably, the particle size of the coarsely divided mushrooms may be between 0.5 and 2 mm. Preferably, the particle size of the coarse mushrooms may be 0.5 mm, 0.6 mm, 0.7 mm, 0.8 mm, 0.9 mm, 1 mm, 1.1 mm, 1.2 mm, 1.3 mm, 1.4 mm, 1.5 mm, 1.6 mm , 1.7 mm, 1.8 mm, 1.9 mm, 2 mm or any value and range covered by any two of the foregoing values.

In this embodiment, in the mushroom residue mixing step (step S05), the mushroom residue and deionized water may be mixed in a mass-volume ratio of 1:20 to 1:250, and then the concentration is 0.05 to 1% cellulose hydrolyzing enzyme is mixed evenly. Preferably, the mushroom residue and deionized water can be mixed in a mass-volume ratio of 1:50 to 1:200, and then a cellulose hydrolyzing enzyme with a concentration of 0.1 to 0.6% is added for uniform mixing. Preferably, the mushroom residue and deionized water can be in a mass-volume ratio of 1:50, 1:55, 1:60, 1:65, 1:70, 1:75, 1:80, 1:85 , 1:90, 1:95, 1:100, 1:105, 1:110, 1:115, 1:120, 1:125, 1:130, 1:135, 1:140, 1:145, 1 : 150, 1: 155, 1: 160, 1: 165, 1: 170, 1: 175, 1: 180, 1: 185, 1: 190, 1: 195, 1: 200 or between any two of the foregoing Mix the proportions of any value and range covered, and then add fibers with a concentration of 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6% or any value and range covered by any two of the foregoing values The hydrolyzed enzyme is evenly mixed.

In this embodiment, the mushroom extract obtained in the foregoing embodiment is rich in β- glucan and triterpenes.

Range: Throughout the present invention, various embodiments of the present invention may be presented in the form of a range. It should be understood that the description in range format is only for convenience and brevity, and should not be construed as a limitation on the scope of the present invention. Therefore, the description of a range should be regarded as specifically disclosing all possible subranges and a single value within the range. For example, the description of the range from 1 to 6 should be considered as having specific disclosed sub-ranges, such as from 1 to 3, 1 to 4, 1 to 5, 2 to 4, 2 to 6, 3 to 6, etc., and in Single and partial numbers in this range, such as 1, 2, 2.7, 3, 4, 5, 5.3, and 6. The foregoing rules apply regardless of the extent of the scope.

The following experimental examples are used to illustrate the method for extracting active ingredients from mushrooms of the present invention, which can achieve the effect of significantly improving the active ingredients in mushroom extracts.

Determination of active ingredients ( β - glucan, triterpene ): refer to the method described in BV McCleary et al. in Measurement of β- Glucan in Mushrooms and Mycelial Products. J AOAC Int., 99(2):364-373 Determination of β -glucan; refer to the determination of triterpenoids in Ganoderma lucidum fermented mycelium by Wang Wei et al., Chinese Edible Fungi, Volume 25, Issue 1, p30-32 for determination of triterpenes.

Determination of total dextran:

(1) Weigh a 100 mg mushroom sample into a test tube.

(2) Add 1.5 mL of 12 M concentrated hydrochloric acid, lock the cap of the test tube, and stir thoroughly.

(3) Put the test tube in the cold water bath for 2 hours, and stir several times to ensure that the β -glucan is completely dissolved.

(4) Add 10 mL of deionized water to each test tube, cover the test tube, and stir for 20 seconds.

(5) Place the test tube in a boiling water bath (about 100°C) and react for 2 hours.

(6) Take out the test tube and cool it to room temperature, and then carefully loosen the test tube cover.

(7) Use 200 mM sodium acetate buffer (pH 5) to quantitatively transfer the contents of each test tube to a 100 mL volumetric flask.

(8) Add 10 mL of 2 M potassium hydroxide to the volumetric flask, and adjust the volume with 200 mM sodium acetate buffer (pH 5). The contents were mixed thoroughly by inversion, centrifuged at 13000 rpm for 5 minutes, and the supernatant was collected as the test substance.

(9) Take 0.1 mL of the object to be tested and put it into a glass test tube.

(10) Add 0.1 mL exo-1,3- β -glucanase (100 U/mL) and β -glucosidase (4 U/mL) to each test tube, and 200 mM sodium acetate buffer ( pH 5) The mixture is reacted at 40°C for 60 minutes.

(11) Add 3 mL of glucose oxidase/peroxidase mixture (GOPOD) to each test tube, and react at 40°C for 20 minutes.

(12) Measure the absorbance at a wavelength of 510 nm with a spectrophotometer.

(13) Calculate the total glucan concentration: the calculation formula is as follows

Total dextran concentration (%, w/w) = (absorbance value of analyte-absorbance value of blank group) * (100 / D-glucose standard absorbance value) / weight of analyte * 90

α - Dextran Determination:

(1) Take 0.1 mL of the test substance and put it in a test tube.

(2) Put a small magnetic rod into each test tube, add 2 mL of 2 M potassium hydroxide, and stir in an ice water bath. During stirring, add 8 mL 1.2 M sodium acetate buffer (pH 3.8), 0.2 mL amyloglucosidase (1630 U/mL) and invertase (500 U/mL), and react for 20 minutes.

(3) Place the test tube in a 40°C water bath and react for 30 minutes.

(4) Centrifuge at 13000 rpm for 3 minutes, and collect the supernatant.

(5) Put 0.1 mL of the supernatant into a test tube, add 0.1 mL of 200 mM sodium acetate buffer (pH 5) and 3 mL of GOPOD reagent, and react at 40°C for 20 minutes.

(6) Measure the absorbance at a wavelength of 510 nm with a spectrophotometer.

(7) Calculate the concentration of α-glucan: the calculation formula is as follows

When the final volume is 100 mL, α-glucan concentration (%, w/w) = (absorbance value of the analyte-absorbance value of the blank group) * (100 / D-glucose standard absorbance value) / weight of the analyte * 90

When the final volume is 10.3 mL, the concentration of α-glucan (%, w/w) = (absorbance value of the analyte-absorbance value of the blank group) * (100 / D-glucose standard absorbance value) / weight of the analyte * 9.27

Calculation of β - glucan content: Measure the concentration of total glucan and α-glucan by the above method, and then subtract α-glucan from the total glucan to obtain the calculated value of β-glucan. Calculated as follows:

β -glucan concentration = (total glucan concentration)-(α-glucan concentration)

Determination of triterpene content:

(1) Take 1 mL of standard oleanolic acid (Oleanolic acid) of different concentrations or the test substance of mushrooms, put it in a test tube, place it on a water bath at 100°C, evaporate to dryness, and put it at room temperature.

(2) Add 0.3 mL of 5% vanillin-glacial acetic acid and 1 mL perchloric acid, and mix well.

(3) Heat it in a water bath at 70°C for 25 minutes, then move it to an ice water bath to cool for 3 minutes, take it out and put it at room temperature.

(4) Add 10 mL of glacial acetic acid, mix well, and place at room temperature to react for 15 minutes.

(5) Measure the absorbance of different concentrations of oleanolic acid (Oleanolic acid) standards or mushrooms to be tested with a spectrophotometer at 550 nm. And draw the triterpene standard curve with different concentrations of oleanolic acid (see Figure 3), and substitute the absorbance value of the mushroom test substance into the linear regression equation of the triterpene standard curve to calculate the mushroom test substance The content of triterpenes.

The following are different experimental examples of extracting active ingredients from Brazilian mushrooms using the method for extracting active ingredients from mushrooms of the present invention:

Experimental example 1:

The Brazilian mushroom fruit bodies are crushed with a coarse crusher to obtain coarse Brazilian mushrooms. Then, the Brazilian mushroom coarse crushed materials and deionized water are mixed uniformly in a mass-volume ratio of 3:400 to obtain coarse crushed materials. mixture. Then, the coarsely crushed mixture is added to the subcritical extraction tank for extraction. The pressure of the subcritical extraction tank is set to 1500 psi, the extraction temperature is 100°C, and the extraction time is 10 minutes. After the extraction, the crude Brazilian mushroom extract is obtained. Centrifuge the crude Brazilian mushroom extract at 6000 rpm for 20 minutes, collect the supernatant separated from the crude Brazilian mushroom extract to obtain the first supernatant, and collect the lower sediment separated from the crude Brazilian mushroom extract. To get Brazilian mushroom residue. Next, the Brazilian mushroom residue and deionized water are mixed uniformly in a mass-volume ratio of 1:50, and then 0.1% of the cellulose hydrolase concentration is added for uniform mixing to obtain a reaction mixture. The reaction mixture is added to a high-pressure hydrolysis tank for extraction. The pressure of the high-pressure hydrolysis tank is set to 1000 psi, the extraction temperature is 30°C, and the extraction time is 30 minutes. After the extraction, the hydrolyzate is obtained. Then, the hydrolyzed liquid was centrifuged at 6000 rpm for 20 minutes, the supernatant liquid separated from the hydrolyzed liquid was collected to obtain the second supernatant liquid, and the precipitated residue was removed. Finally, the first supernatant liquid and the second supernatant liquid are mixed, and concentrated under reduced pressure at 37° C. to twice the weight of the raw material (w/w) to obtain the Brazilian mushroom extract.

Experimental example 2:

The Brazilian mushroom fruit bodies are crushed with a coarse crusher to obtain coarse Brazilian mushrooms. Then, the Brazilian mushroom coarse crushed materials and deionized water are mixed uniformly in a mass-volume ratio of 9:400 to obtain coarse crushed materials. mixture. Then, the coarsely crushed mixture is added to the subcritical extraction tank for extraction. The pressure of the subcritical extraction tank is set to 1500 psi, the extraction temperature is 150° C., and the extraction time is 30 minutes. After the extraction is completed, a crude extract of Brazilian mushrooms is obtained. Centrifuge the crude Brazilian mushroom extract at 6000 rpm for 20 minutes, collect the supernatant separated from the crude Brazilian mushroom extract to obtain the first supernatant, and collect the lower sediment separated from the crude Brazilian mushroom extract. To get Brazilian mushroom residue. Then, the Brazilian mushroom residue and deionized water are mixed uniformly in a mass-volume ratio of 1:50, and then 0.6% of the cellulose hydrolase concentration is added for uniform mixing to obtain a reaction mixture. The reaction mixture is added to a high-pressure hydrolysis tank for extraction. The pressure of the high-pressure hydrolysis tank is set to 1000 psi, the extraction temperature is 60°C, and the extraction time is 90 minutes. After the extraction, the hydrolyzate is obtained. Then, the hydrolyzed liquid was centrifuged at 6000 rpm for 20 minutes, the supernatant liquid separated from the hydrolyzed liquid was collected to obtain the second supernatant liquid, and the precipitated residue was removed. Finally, the first supernatant liquid and the second supernatant liquid are mixed, and concentrated under reduced pressure at 37° C. to twice the weight of the raw material (w/w) to obtain the Brazilian mushroom extract.

Experimental example 3:

The Brazilian mushroom fruit bodies are crushed with a coarse crusher to obtain coarse Brazilian mushrooms. Then, the Brazilian mushroom coarse crushed materials and deionized water are mixed uniformly in a mass-volume ratio of 6:400 to obtain coarse crushed materials. mixture. Then, the coarsely crushed mixture is added to the subcritical extraction tank for extraction, the pressure of the subcritical extraction tank is set to 3000 psi, the extraction temperature is 150° C., and the extraction time is 10 minutes. After the extraction is completed, a crude extract of Brazilian mushrooms is obtained. Centrifuge the crude Brazilian mushroom extract at 6000 rpm for 20 minutes, collect the supernatant separated from the crude Brazilian mushroom extract to obtain the first supernatant, and collect the lower sediment separated from the crude Brazilian mushroom extract. To get Brazilian mushroom residue. Then, the Brazilian mushroom residue and deionized water are mixed uniformly in a mass-volume ratio of 1:50, and then 0.3% of the cellulose hydrolase concentration is added for uniform mixing to obtain a reaction mixture. The reaction mixture is added to a high-pressure hydrolysis tank for extraction. The pressure of the high-pressure hydrolysis tank is set to 2000 psi, the extraction temperature is 60°C, and the extraction time is 30 minutes. After the extraction, the hydrolyzate is obtained. Then, the hydrolyzed liquid was centrifuged at 6000 rpm for 20 minutes, the supernatant liquid separated from the hydrolyzed liquid was collected to obtain the second supernatant liquid, and the precipitated residue was removed. Finally, the first supernatant liquid and the second supernatant liquid are mixed, and concentrated under reduced pressure at 37° C. to twice the weight of the raw material (w/w) to obtain the Brazilian mushroom extract.

Experimental example 4:

The Brazilian mushroom fruit bodies are crushed with a coarse crusher to obtain coarse Brazilian mushrooms. Then, the Brazilian mushroom coarse crushed materials and deionized water are mixed uniformly in a mass-volume ratio of 9:400 to obtain coarse crushed materials. mixture. Then, the coarsely crushed mixture is added to the subcritical extraction tank for extraction, the pressure of the subcritical extraction tank is set to 3000 psi, the extraction temperature is 100°C, the extraction time is 20 minutes, and the extraction is completed to obtain the crude Brazilian mushroom extract. Centrifuge the crude Brazilian mushroom extract at 6000 rpm for 20 minutes, collect the supernatant separated from the crude Brazilian mushroom extract to obtain the first supernatant, and collect the lower sediment separated from the crude Brazilian mushroom extract. To get Brazilian mushroom residue. Then, the Brazilian mushroom residue and deionized water are mixed uniformly in a mass-volume ratio of 1:50, and then 0.6% of the cellulose hydrolase concentration is added for uniform mixing to obtain a reaction mixture. The reaction mixture is added to a high-pressure hydrolysis tank for extraction. The pressure of the high-pressure hydrolysis tank is set to 2000 psi, the extraction temperature is 30°C, and the extraction time is 60 minutes. After the extraction, the hydrolyzate is obtained. Then, the hydrolyzed liquid was centrifuged at 6000 rpm for 20 minutes, the supernatant liquid separated from the hydrolyzed liquid was collected to obtain the second supernatant liquid, and the precipitated residue was removed. Finally, the first supernatant liquid and the second supernatant liquid are mixed, and concentrated under reduced pressure at 37° C. to twice the weight of the raw material (w/w) to obtain the Brazilian mushroom extract.

Experimental example 5:

The Brazilian mushroom fruit bodies are crushed with a coarse crusher to obtain coarse Brazilian mushrooms. Then, the Brazilian mushroom coarse crushed materials and deionized water are mixed uniformly in a mass-volume ratio of 3:400 to obtain coarse crushed materials. mixture. Then, the coarsely crushed mixture is added to the subcritical extraction tank for extraction. The pressure of the subcritical extraction tank is set to 4500 psi, the extraction temperature is 150°C, and the extraction time is 20 minutes. After the extraction is completed, a crude extract of Brazilian mushrooms is obtained. Centrifuge the crude Brazilian mushroom extract at 6000 rpm for 20 minutes, collect the supernatant separated from the crude Brazilian mushroom extract to obtain the first supernatant, and collect the lower sediment separated from the crude Brazilian mushroom extract. To get Brazilian mushroom residue. Next, the Brazilian mushroom residue and deionized water are mixed uniformly in a mass-volume ratio of 1:50, and then 0.1% of the cellulose hydrolase concentration is added for uniform mixing to obtain a reaction mixture. The reaction mixture is added to a high-pressure hydrolysis tank for extraction. The pressure of the high-pressure hydrolysis tank is set to 3000 psi, the extraction temperature is 60°C, and the extraction time is 60 minutes. After the extraction, the hydrolyzate is obtained. Then, the hydrolyzed liquid was centrifuged at 6000 rpm for 20 minutes, the supernatant liquid separated from the hydrolyzed liquid was collected to obtain the second supernatant liquid, and the precipitated residue was removed. Finally, the first supernatant liquid and the second supernatant liquid are mixed, and concentrated under reduced pressure at 37° C. to twice the weight of the raw material (w/w) to obtain the Brazilian mushroom extract.

Experimental example 6:

The Brazilian mushroom fruit bodies are crushed with a coarse crusher to obtain coarse Brazilian mushrooms. Then, the Brazilian mushroom coarse crushed materials and deionized water are mixed uniformly in a mass-volume ratio of 6:400 to obtain coarse crushed materials. mixture. Then, the coarsely crushed mixture is added to the subcritical extraction tank for extraction, the pressure of the subcritical extraction tank is set to 4500 psi, the extraction temperature is 100°C, the extraction time is 30 minutes, and the extraction is completed to obtain the crude Brazilian mushroom extract. Centrifuge the crude Brazilian mushroom extract at 6000 rpm for 20 minutes, collect the supernatant separated from the crude Brazilian mushroom extract to obtain the first supernatant, and collect the lower sediment separated from the crude Brazilian mushroom extract. To get Brazilian mushroom residue. Then, the Brazilian mushroom residue and deionized water are mixed uniformly in a mass-volume ratio of 1:50, and then 0.3% of the cellulose hydrolase concentration is added for uniform mixing to obtain a reaction mixture. The reaction mixture is added to a high-pressure hydrolysis tank for extraction. The pressure of the high-pressure hydrolysis tank is set to 3000 psi, the extraction temperature is 30°C, and the extraction time is 90 minutes. After the extraction, the hydrolyzate is obtained. Then, the hydrolyzed liquid was centrifuged at 6000 rpm for 20 minutes, the supernatant liquid separated from the hydrolyzed liquid was collected to obtain the second supernatant liquid, and the precipitated residue was removed. Finally, the first supernatant liquid and the second supernatant liquid are mixed, and concentrated under reduced pressure at 37° C. to twice the weight of the raw material (w/w) to obtain the Brazilian mushroom extract.

Experimental example 7:

The Brazilian mushroom fruit bodies are crushed with a coarse crusher to obtain coarse Brazilian mushrooms. Then, the Brazilian mushroom coarse crushed materials and deionized water are mixed uniformly in a mass-volume ratio of 9:400 to obtain coarse crushed materials. mixture. Then, the coarsely crushed mixture is added to the subcritical extraction tank for extraction, the pressure of the subcritical extraction tank is set to 4500 psi, the extraction temperature is 125°C, and the extraction time is 10 minutes. After the extraction is completed, a crude extract of Brazilian mushrooms is obtained. Centrifuge the crude Brazilian mushroom extract at 6000 rpm for 20 minutes, collect the supernatant separated from the crude Brazilian mushroom extract to obtain the first supernatant, and collect the lower sediment separated from the crude Brazilian mushroom extract. To get Brazilian mushroom residue. Then, the Brazilian mushroom residue and deionized water are mixed uniformly in a mass-volume ratio of 1:50, and then 0.6% of the cellulose hydrolase concentration is added for uniform mixing to obtain a reaction mixture. The reaction mixture is added to a high-pressure hydrolysis tank for extraction. The pressure of the high-pressure hydrolysis tank is set to 3000 psi, the extraction temperature is 45° C., and the extraction time is 30 minutes. After the extraction, the hydrolyzate is obtained. Then, the hydrolyzed liquid was centrifuged at 6000 rpm for 20 minutes, the supernatant liquid separated from the hydrolyzed liquid was collected to obtain the second supernatant liquid, and the precipitated residue was removed. Finally, the first supernatant liquid and the second supernatant liquid are mixed, and concentrated under reduced pressure at 37° C. to twice the weight of the raw material (w/w) to obtain the Brazilian mushroom extract.

Experimental example 8:

The Brazilian mushroom fruit bodies are crushed with a coarse crusher to obtain coarse Brazilian mushrooms. Then, the Brazilian mushroom coarse crushed materials and deionized water are mixed uniformly in a mass-volume ratio of 3:400 to obtain coarse crushed materials. mixture. Then, the coarsely crushed mixture is added to the subcritical extraction tank for extraction, the pressure of the subcritical extraction tank is set to 3000 psi, the extraction temperature is 100°C, the extraction time is 30 minutes, and the extraction is completed to obtain the crude Brazilian mushroom extract. Centrifuge the crude Brazilian mushroom extract at 6000 rpm for 20 minutes, collect the supernatant separated from the crude Brazilian mushroom extract to obtain the first supernatant, and collect the lower sediment separated from the crude Brazilian mushroom extract. To get Brazilian mushroom residue. Then, the Brazilian mushroom residue and deionized water are mixed uniformly in a mass-volume ratio of 1:50, and then 0.3% of the cellulose hydrolase concentration is added for uniform mixing to obtain a reaction mixture. The reaction mixture was added to a high-pressure hydrolysis tank for extraction. The pressure of the high-pressure hydrolysis tank was set to 3000 psi, the extraction temperature was 30°C, and the extraction time was 60 minutes. After the extraction, the hydrolyzate was obtained. Then, the hydrolyzed liquid was centrifuged at 6000 rpm for 20 minutes, the supernatant liquid separated from the hydrolyzed liquid was collected to obtain the second supernatant liquid, and the precipitated residue was removed. Finally, the first supernatant liquid and the second supernatant liquid are mixed, and concentrated under reduced pressure at 37° C. to twice the weight of the raw material (w/w) to obtain the Brazilian mushroom extract.

The Brazilian mushroom extracts obtained in the above experimental examples 1 to 8 were tested for the content of β -glucan and triterpenes according to the above method, and the test results are as follows in Table 1.

Table 1: Brazilian Mushroom Extract β- glucan content (%) Triterpene content (%) Experimental example 1 3.92±0.21 0.43±0.04 Experimental example 2 3.83±0.05 0.79±0.09 Experimental example 3 3.05±0.18 0.65±0.03 Experimental example 4 3.01±0.06 0.51±0.04 Experimental example 5 2.42±0.27 0.83±0.16 Experimental example 6 3.25±0.16 0.58±0.07 Experimental example 7 2.05±0.05 0.57±0.11 Experimental example 8 4.16±0.22 0.53±0.08

As shown in Table 1 above, the Brazilian mushroom extract obtained by the extraction method of the mushroom active ingredients of the present invention is rich in β- glucan and triterpenes. In the total amount of Brazilian mushroom extract, the content of β- glucan can reach at least 2%, and the content of triterpenes can reach at least 0.39%.

Although the above experimental examples take the extraction of Brazilian mushrooms as an example for illustration, they are not a limitation of the present invention. The method for extracting active ingredients from mushrooms of the present invention can also be used to extract Phellinus igniarius, Tremella, or other common knowledge in the technical field. Known mushrooms.

In summary, the method for extracting active ingredients from mushrooms of the present invention can achieve the effect of increasing the yield of active ingredients in mushroom extracts, and can avoid traditional extraction methods with long boiling time, high temperature, and low yield. And other problems, it is not necessary to use a large amount of organic solvents.

The above description is only illustrative, and not restrictive. Any equivalent modification or alteration that does not deviate from the spirit and scope of the present invention shall be included in the scope of the appended patent application.

S01, S02, S03, S04, S05, S06, S07, S08: steps

Fig. 1 is a schematic diagram of a preferred embodiment of the method for extracting active ingredients from mushrooms of the present invention. Fig. 2 is a flow chart of a preferred embodiment of the method for extracting active ingredients from mushrooms of the present invention. Figure 3 shows the standard curve of triterpene.

S01, S02, S03, S04, S05, S06, S07, S08: steps

Claims (14)

A method for extracting active ingredients from mushrooms, including the following steps: In a crushing step, a mushroom is crushed by a coarse crusher to obtain a coarse mushroom; In a mixing step, the crude mushrooms and deionized water are mixed uniformly in a mass-volume ratio of 1:400 to 10:400 to obtain a coarsely crushed mixture; In a subcritical extraction step, the crude mixture is added to a subcritical extraction tank for extraction. The pressure of the subcritical extraction tank is between 1000 and 5000 psi, and the extraction temperature is between 50 and 200°C. The time is between 5 and 40 minutes, after the extraction, a crude extract of mushrooms is obtained; In a first centrifugation step, the crude mushroom extract is centrifuged at 5000 rpm to 15000 rpm for 5 to 30 minutes, and the supernatant separated from the crude mushroom extract is collected to obtain a first supernatant And collect the lower sediment separated from the crude extract of the mushrooms to obtain a mushroom residue; A mushroom residue mixing step, uniformly mixing the mushroom residue with a deionized water and a cellulose hydrolyzing enzyme to obtain a reaction mixture; In a high-pressure hydrolysis step, the reaction mixture is added to a high-pressure hydrolysis tank for extraction. The pressure of the high-pressure hydrolysis tank is between 500 and 5000 psi, the extraction temperature is between 10 and 100°C, and the extraction time is between 10 To 100 minutes, a hydrolysate is obtained after the extraction is completed; In a second centrifugation step, the hydrolyzed liquid is centrifuged at a rotation speed of 5000 rpm to 15000 rpm for 5 to 30 minutes, and the supernatant separated from the hydrolyzed liquid is collected to obtain a second supernatant; and In a vacuum concentration step, the first supernatant and the second supernatant are mixed, and concentrated under reduced pressure at 37° C. to obtain a mushroom extract. The extraction method according to claim 1, wherein in the mixing step, the mass-volume ratio of the crude mushrooms to the deionized water is between 3:400 and 9:400. The extraction method according to claim 1, wherein in the subcritical extraction step, the pressure of the subcritical extraction tank is between 1500 and 4500 psi, the extraction temperature is between 100 and 150°C, and the extraction time is between Between 10 and 30 minutes. The extraction method according to claim 1, wherein in the high-pressure hydrolysis step, the pressure of the high-pressure hydrolysis tank is between 1000 and 3000 psi, the extraction temperature is between 30 and 60°C, and the extraction time is between 30 and Between 90 minutes. The extraction method according to claim 1, wherein in the first centrifugation step, the crude mushroom extract is centrifuged at a rotation speed of 6000 rpm to 13000 rpm for 10 to 20 minutes. The extraction method according to claim 1, wherein in the second centrifugation step, the hydrolyzed liquid is centrifuged at a rotation speed of 6000 rpm to 13000 rpm for 10 to 20 minutes. The extraction method according to claim 1, wherein in the crushing step, the mushrooms are crushed with a coarse crusher for 10 to 50 minutes. The extraction method according to claim 7, wherein in the crushing step, the mushrooms are crushed with a coarse crusher for 20 to 30 minutes. The extraction method according to claim 1, wherein the particle size of the crude mushrooms is between 0.1 and 3 mm. The extraction method according to claim 9, wherein the size of the coarsely divided mushrooms is between 0.5 and 2 mm. The extraction method according to claim 1, wherein in the mushroom residue mixing step, the mushroom residue and the deionized water are mixed in a mass-volume ratio of 1:20 to 1:250, and then the concentration is added 0.05 to 1% of the cellulose hydrolase is uniformly mixed. The extraction method according to claim 11, wherein in the mushroom residue mixing step, the mushroom residue and the deionized water are mixed in a mass-volume ratio of 1:50 to 1:200, and then the concentration is added 0.1 to 0.6% of the cellulose hydrolyzing enzyme is uniformly mixed. The extraction method according to claim 1, wherein the mushroom extract is rich in β- glucan and triterpenes. The extraction method according to claim 1, wherein the mushroom is Brazilian mushroom, Phellinus igniarius or Tremella.

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