Low-temperature efficient extraction method of tricholoma matsutake active ingredients

Abstract

The invention relates to the technical field of food processing, and discloses a low-temperature efficient extraction method of tricholoma matsutake active ingredients, which comprises the following steps: S1, antioxidant pretreatment; s2, vacuum ultrasonic synergistic extraction: transferring the tricholoma matsutake slices soaked in the step S1 to vacuum ultrasonic extraction equipment, synchronously performing drying and active ingredient extraction under low-temperature and vacuum conditions in combination with ultrasonic assistance, and collecting tricholoma matsutake small molecular group cell stock solution in real time; s3, gradient interception solid-liquid separation; s4, modified membrane separation and purification; s5, performing falling film concentration under nitrogen protection; and S6, performing pressure gradient non-thermal sterilization. According to the method, the technical scheme of combining antioxidant protection of composite pretreatment liquid with vacuum ultrasonic synergistic extraction is adopted, so that the technical effect of efficiently retaining active ingredients in tricholoma matsutake is achieved; the defects that the active components are easily degraded or structurally damaged due to oxidation and mechanical action, and the retention effect is poor are overcome.

Description

Technical Field

[0001] This invention relates to the field of food processing technology, specifically to a low-temperature, high-efficiency extraction method for active ingredients in matsutake mushrooms. Background Technology

[0002] Matsutake mushrooms are a precious edible and medicinal fungus in my country, rich in various active ingredients such as matsutake alcohol, polypeptides, and polysaccharides. These ingredients possess multiple physiological functions, including antioxidant, immunomodulatory, and anti-inflammatory effects, and have broad application prospects in the food, health product, and pharmaceutical fields. With the increasing market demand for natural active ingredients, the efficient extraction and preservation of these active ingredients from matsutake mushrooms has become one of the core research directions in the field of matsutake deep processing. The extraction effect directly affects the functional value and market competitiveness of the product.

[0003] Existing extraction technologies for active ingredients in matsutake mushrooms often employ traditional extraction methods such as single-solvent immersion or conventional ultrasonic extraction, which lack targeted antioxidant protection designs. Furthermore, active ingredients are prone to oxidative degradation upon contact with air during the extraction process. In addition, the continuous mechanical action of conventional ultrasonic extraction can damage the molecular structure of active ingredients, further resulting in poor retention of active ingredients. This makes it difficult to fully realize the resource value of matsutake mushrooms and limits the quality improvement and industrialization of related products. Summary of the Invention

[0004] To address the shortcomings of existing technologies, this invention provides a low-temperature, high-efficiency extraction method for active ingredients of matsutake mushrooms. This method solves the problem that existing technologies, such as simple two-stage filtration or ordinary polyethersulfone membrane separation, result in incomplete removal of impurities from the active ingredients of matsutake mushrooms, leaving behind a large amount of protein, colloid, and other impurities, which affects product quality.

[0005] To achieve the above objectives, the present invention provides the following technical solution: a low-temperature, high-efficiency extraction method for active ingredients of matsutake mushrooms, comprising the following steps: S1. Antioxidant pretreatment: Select qualified fresh matsutake mushrooms, wash them with reverse osmosis purified water, drain them in a sterile environment, and then slice them to obtain matsutake slices. Place the matsutake slices in a sterile buffer solution containing antioxidants and soak them at low temperature to maintain the stability of the matsutake cell structure. S2, Vacuum-ultrasonic synergistic extraction: The matsutake slices soaked in S1 are transferred to a vacuum ultrasonic extraction device. Under low temperature and vacuum conditions, combined with ultrasonic assistance, drying and active ingredient extraction are carried out simultaneously, and the matsutake small molecular cluster cell original solution is collected in real time. S3. Gradient retention solid-liquid separation: First, a large-pore coarse filtration membrane is used to remove large particles of dietary fiber, then a fine filtration membrane is used to retain starch particles, and finally a ceramic microfiltration membrane is used to retain colloidal impurities. The three-stage filtration process yields a highly clear filtrate. S4. Modified membrane separation and purification: The filtrate from the three-stage filtration in S3 is passed into a surface-modified ultrafiltration membrane system to selectively retain large protein molecules while simultaneously retaining small molecule active ingredients, and the filtrate rich in matsutake alcohol, peptides and polysaccharides is collected. S5. Nitrogen-protected falling film concentration: The filtrate from S4 is concentrated under low temperature and vacuum conditions, with nitrogen gas introduced throughout the process to isolate oxygen and prevent the active ingredients from oxidizing and degrading, resulting in a highly active matsutake mushroom concentrate. S6. Pressure gradient non-thermal sterilization: The concentrated stock solution of S5 is treated with a three-stage pressure gradient to maximize the retention of active ingredients while ensuring the sterilization effect, so as to obtain high-purity matsutake mushroom extract.

[0006] Preferably, in step S1, the thickness of the matsutake mushroom slices is 2.5-4.5 mm; The qualified fresh matsutake mushrooms contain 0.14% to 0.21% matsutake alcohol, the caps are not fully open and the diameter is 4.5 to 6.5 cm, the stem length is 9 to 14 cm and the diameter is 2.3 to 2.9 cm, and the total time from picking to S1 pretreatment is 3.5 to 9.5 hours. The antioxidant is 0.01-0.03% ascorbate palmitate, which is mixed with phosphate buffer to form a composite pretreatment solution; The pH of the composite pretreatment solution was 6.4-6.9, the mass-to-volume ratio of the solution to the matsutake slices was 1:2.11:3.1 g / mL, the soaking temperature was 15℃, the soaking time was 12-18 minutes, and the difference between the buffer solution temperature and the fresh matsutake transport temperature was 0.5-2.5℃.

[0007] Preferably, in step S1, the reverse osmosis purified water cleaning water pressure is 0.3-0.4 MPa, the cleaning is performed twice, and the cleaning time for each cleaning is 35-45 seconds. After draining, the surface moisture content of the matsutake mushroom slices was 6%-14%.

[0008] Preferably, in step S2, the low temperature condition is 17°C and the vacuum condition is a vacuum degree of 0.05-0.07 MPa; Ultrasound-assisted parameters include: frequency 2040kHz, power density 0.3-0.5W / cm². 2 The ultrasound is activated intermittently. The heating rate of the vacuum extraction equipment is 0.6-0.9℃ / 10 minutes, the total drying and extraction time is 3.2-4.8 hours, the original solution collected simultaneously accounts for 71%-74% of the total cell sap content of matsutake mushrooms, and the start-stop time difference between original solution collection and drying and extraction is 1.5-4.5 minutes, which increases the dissolution rate of active ingredients by 15%-20%.

[0009] Preferably, in step S2, the collected stock solution is temporarily stored in a sterile, sealed container, and the storage conditions include: Temperature 13℃, pressure 0.022-0.028MPa, temporary storage time 1.5-3.5 hours, immediately proceed to S3 gradient retention filtration step; A small amount of nitrogen gas was continuously introduced during the temporary storage process at a flow rate of 510 mL / min.

[0010] Preferably, in step S3, the gradient filtration includes: a coarse filter membrane with a pore size of 0.5 μm, an operating pressure of 0.16-0.24 MPa, and a flow rate of 3.1-3.9 m / s². 3 / h; The fine filtration membrane has a pore size of 0.2 μm, an operating pressure of 0.21-0.29 MPa, and a flow rate of 2.1-2.9 m / s. 3 / h; The ceramic microfiltration membrane has a pore size of 50-80 nm, an operating pressure of 0.3-0.4 MPa, and a flow rate of 1.5-2.0 m / s. 3 / h; During the three-stage filtration process, the filtrate temperature is maintained at 15℃, which is consistent with the collection temperature of the S2 stock solution, and the impurity removal rate is 99.2%.

[0011] Preferably, in step S4, the surface-modified ultrafiltration membrane is a polyethersulfone and graphene composite modified membrane with a pore size of 1119 nm and a molecular weight cutoff of 1129 kDa for protein impurities. The transmembrane pressure difference of the ultrafiltration membrane system was controlled at 0.04-0.09 MPa. Cross-flow filtration was adopted with a cross-flow velocity of 11.5 m / s. The total content of matsutake alcohol, matsutake polypeptide and matsutake polysaccharide in the filtrate was 0.18%-0.29%, the protein retention rate was 97.2%-98.8%, and the permeation rate of active ingredients was increased by 12%-18%.

[0012] Preferably, in step S5, the temperature of vacuum falling film concentration is 3742°C and the vacuum degree is 0.072-0.088 MPa; The nitrogen protection flow rate is 2030 mL / min, covering the surface of the concentrate to form an inert atmosphere; The concentration of the concentrated matsutake mushroom concentrate was 8.5-11.5 Brix, the loss rate of active ingredients during the concentration process was 1.2%-4.8%, and the viscosity at 25℃ was 1624 mPas.

[0013] Preferably, in step S6, the three-stage pressure gradient comprises three phases: low-pressure pre-sterilization, medium-pressure stabilization, and high-pressure inactivation: Low-pressure pre-sterilization: 180-280 MPa, processing time: 1 minute; Medium-pressure stable pressure 300-400MPa, processing time 30 seconds; High-pressure inactivation: 420-530 MPa, holding time: 3 minutes; The entire process is carried out at a temperature of 119℃, using a step-by-step pressurization method.

[0014] Preferably, in step S2, the vacuum ultrasonic synergistic extraction further includes: the matsutake mushroom slices are spread to a thickness of 12 cm in the vacuum ultrasonic extraction device; Nitrogen purging was performed every 30 minutes during the extraction process, with a purging time of 12 minutes. After purging, a vacuum of 0.05-0.07 MPa was maintained, and the ultrasonic power was dynamically adjusted according to the extraction time, maintaining a power density of 0.3-0.4 W / cm³ for the first 2 hours. 2 After 1.2 to 2.8 hours, the concentration was adjusted to 0.4 to 0.5 W / cm². 2 .

[0015] This invention provides a low-temperature, high-efficiency extraction method for the active ingredients of matsutake mushrooms. It has the following beneficial effects: 1. This invention achieves the technical effect of efficiently preserving the active ingredients in matsutake mushrooms by adopting a technical solution that combines antioxidant protection of composite pretreatment liquid with vacuum ultrasonic synergistic extraction. Compared with the existing technology of single solvent soaking or ordinary ultrasonic extraction, it solves the shortcomings of the active ingredients being easily degraded or structurally damaged due to oxidation and mechanical action, resulting in poor retention effect.

[0016] 2. This invention employs a synergistic technical solution of gradient retention three-stage filtration and separation using a polyethersulfone and graphene composite modified membrane, which significantly improves the purity of the extracted product. Compared with the existing simple two-stage filtration or ordinary polyethersulfone membrane separation technology, this invention solves the problem of incomplete impurity removal and the presence of a large amount of protein, colloid and other impurities that affect product quality.

[0017] 3. The present invention adopts a technical solution that combines nitrogen-protected falling film concentration with pressure gradient non-thermal sterilization, which achieves the technical effect of ensuring product storage stability and sterilization safety. Compared with the existing technology of vacuum concentration without protection or single pressure sterilization, it solves the shortcomings of active ingredients being easily oxidized and deteriorated during the concentration process, and the components being easily damaged by high pressure and insufficient sterilization uniformity during sterilization. Attached Figure Description

[0018] Figure 1 This is a schematic diagram of a low-temperature, high-efficiency extraction method for active ingredients of matsutake mushroom according to the present invention. Detailed Implementation

[0019] The technical solution of the present invention will now be clearly and completely described with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0020] Please see the appendix Figure 1This invention provides a low-temperature, high-efficiency extraction method for active ingredients from matsutake mushrooms, comprising the following steps: S1. Antioxidant pretreatment: Select qualified fresh matsutake mushrooms, wash them with reverse osmosis purified water, drain them in a sterile environment, and then slice them to obtain matsutake slices. Place the matsutake slices in a sterile buffer solution containing antioxidants and soak them at low temperature to maintain the stability of the matsutake cell structure. The thickness of the matsutake slices is 2.5-4.5 mm; The qualified fresh matsutake mushrooms contain 0.14% to 0.21% matsutake alcohol, the caps are not fully open and the diameter is 4.5 to 6.5 cm, the stem length is 9 to 14 cm and the diameter is 2.3 to 2.9 cm, and the total time from picking to S1 pretreatment is 3.5 to 9.5 hours. The antioxidant is 0.01-0.03% ascorbate palmitate, which is mixed with phosphate buffer to form a composite pretreatment solution; The pH of the composite pretreatment solution was 6.4-6.9, the mass-to-volume ratio of the solution to the matsutake slices was 1:2.11:3.1 g / mL, the soaking temperature was 15℃, the soaking time was 12-18 minutes, and the difference between the buffer solution temperature and the fresh matsutake transport temperature was 0.5-2.5℃.

[0021] The reverse osmosis purified water cleaning pressure is 0.3-0.4 MPa, the cleaning is performed twice, and the cleaning time for each cleaning is 35-45 seconds; After draining, the surface moisture content of the matsutake mushroom slices was 6%-14%. S2, Vacuum-ultrasonic synergistic extraction: The matsutake slices soaked in S1 are transferred to a vacuum ultrasonic extraction device. Under low temperature and vacuum conditions, combined with ultrasonic assistance, drying and active ingredient extraction are carried out simultaneously, and the matsutake small molecular cluster cell original solution is collected in real time. The low temperature condition is 17℃, and the vacuum condition is a vacuum degree of 0.05-0.07MPa. Ultrasound-assisted parameters include: frequency 2040kHz, power density 0.3-0.5W / cm². 2 The ultrasound is activated intermittently. The heating rate of the vacuum extraction equipment is 0.6-0.9℃ / 10 minutes, the total drying and extraction time is 3.2-4.8 hours, the original solution collected simultaneously accounts for 71%-74% of the total cell sap content of matsutake mushrooms, and the start-stop time difference between original solution collection and drying and extraction is 1.5-4.5 minutes, which increases the dissolution rate of active ingredients by 15%-20%.

[0022] The collected stock solution is temporarily stored in a sterile, sealed container under the following conditions: Temperature 13℃, pressure 0.022-0.028MPa, temporary storage time 1.5-3.5 hours, immediately proceed to S3 gradient retention filtration step; A small amount of nitrogen gas was continuously introduced during the temporary storage process at a flow rate of 510 mL / min. Vacuum-ultrasonic synergistic extraction also includes: the matsutake mushroom slices are spread to a thickness of 12 cm in the vacuum-ultrasonic extraction equipment; Nitrogen purging was performed every 30 minutes during the extraction process, with a purging time of 12 minutes. After purging, a vacuum of 0.05-0.07 MPa was maintained, and the ultrasonic power was dynamically adjusted according to the extraction time, maintaining a power density of 0.3-0.4 W / cm³ for the first 2 hours. 2 After 1.2 to 2.8 hours, the concentration was adjusted to 0.4 to 0.5 W / cm². 2 ; S3. Gradient retention solid-liquid separation: First, a large-pore coarse filtration membrane is used to remove large particles of dietary fiber, then a fine filtration membrane is used to retain starch particles, and finally a ceramic microfiltration membrane is used to retain colloidal impurities. The three-stage filtration process yields a highly clear filtrate. The gradient retention filtration system includes: a coarse filter membrane with a pore size of 0.5 μm, an operating pressure of 0.16-0.24 MPa, and a flow rate of 3.1-3.9 m / s². 3 / h; The fine filtration membrane has a pore size of 0.2 μm, an operating pressure of 0.21-0.29 MPa, and a flow rate of 2.1-2.9 m / s. 3 / h; The ceramic microfiltration membrane has a pore size of 50-80 nm, an operating pressure of 0.3-0.4 MPa, and a flow rate of 1.5-2.0 m / s. 3 / h; During the three-stage filtration process, the filtrate temperature is maintained at 15℃, which is consistent with the collection temperature of the S2 stock solution, and the impurity removal rate is 99.2%. S4. Modified membrane separation and purification: The filtrate from the three-stage filtration in S3 is passed into a surface-modified ultrafiltration membrane system to selectively retain large protein molecules while simultaneously retaining small molecule active ingredients, and the filtrate rich in matsutake alcohol, peptides and polysaccharides is collected. Among them, the surface-modified ultrafiltration membrane is a polyethersulfone and graphene composite modified membrane with a pore size of 1119 nm and a molecular weight cutoff of 1129 kDa for protein impurities. The transmembrane pressure difference of the ultrafiltration membrane system was controlled at 0.04-0.09 MPa. Cross-flow filtration was adopted with a cross-flow velocity of 11.5 m / s. The total content of matsutake alcohol, matsutake polypeptide and matsutake polysaccharide in the filtrate was 0.18%-0.29%, the protein retention rate was 97.2%-98.8%, and the permeation rate of active ingredients was increased by 12%-18%. S5. Nitrogen-protected falling film concentration: The filtrate from S4 is concentrated under low temperature and vacuum conditions, with nitrogen gas introduced throughout the process to isolate oxygen and prevent the active ingredients from oxidizing and degrading, resulting in a highly active matsutake mushroom concentrate. The vacuum falling film concentration was performed at a temperature of 3742℃ and a vacuum level of 0.072-0.088 MPa. The nitrogen protection flow rate is 2030 mL / min, covering the surface of the concentrate to form an inert atmosphere; The concentration of the concentrated matsutake mushroom concentrate was 8.5-11.5 Brix, the loss rate of active ingredients during the concentration process was 1.2%-4.8%, and the viscosity at 25℃ was 1624 mPas. S6. Pressure gradient non-thermal sterilization: The concentrated stock solution of S5 is treated with a three-stage pressure gradient to maximize the retention of active ingredients while ensuring the sterilization effect, so as to obtain high-purity matsutake extract. The three-stage pressure gradient consists of three phases: low-pressure pre-sterilization, medium-pressure stabilization, and high-pressure inactivation. Low-pressure pre-sterilization: 180-280 MPa, processing time: 1 minute; Medium-pressure stable pressure 300-400MPa, processing time 30 seconds; High-pressure inactivation: 420-530 MPa, holding time: 3 minutes; The entire process is carried out at a temperature of 119℃, using a step-by-step pressurization method.

[0023] The following is a description with reference to specific embodiments: Example 1 S1. Antioxidant pretreatment: Matsutake alcohol content 0.14%, cap diameter 4.5cm, stipe length 9cm, diameter 2.3cm, time from harvesting to pretreatment 3.5 hours, washing water pressure 0.3MPa, washing time 35 seconds / time, drained moisture content 6%, slice thickness 2.5mm, ascorbate palmitate concentration in the compound pretreatment solution 0.01%, pH 6.4, material-to-liquid ratio 1:2.1g / mL, soaking temperature 1℃, time 12 minutes, fresh matsutake transportation temperature 0℃, buffer solution temperature difference from transportation temperature 0.5℃; S2. Vacuum-ultrasonic synergistic extraction: Spreading thickness 1cm, vertical distance between vibrating plate and sheet 3cm, extraction temperature 1℃, vacuum degree 0.05MPa, ultrasonic frequency 20kHz, power density 0.3W / cm³. 2 0.4 W / cm for the first 2 hours 2 After 1.2 hours, the heating rate was 0.6℃ / 10 minutes, the extraction time was 3.2 hours, the original solution collection rate was 71%, the start-stop time difference was 1.5 minutes, the nitrogen purging was 1 minute / 30 minutes, the temporary storage temperature was 1℃, the pressure was 0.022MPa, the time was 1.5 hours, and the nitrogen flow rate was 5mL / min. S3, Gradient retention solid-liquid separation: coarse filtration membrane pore size 0.5 μm, operating pressure 0.16 MPa, flow rate 3.1 m / s². 3 / h, fine filtration membrane pore size 0.2m, operating pressure 0.21MPa, flow rate 2.1m 3 / h, ceramic microfiltration membrane pore size 50nm, operating pressure 0.3MPa, flow rate 1.5m3 / h, filtrate temperature 1℃; S4. Modified membrane separation and purification: Ultrafiltration membrane pore size 11nm, molecular weight cutoff 11kDa, transmembrane pressure difference 0.04MPa, cross-flow velocity 1.0m / s, total content of matsutake alcohol, polypeptides and polysaccharides in filtrate 0.18%, protein rejection rate 97.2%; S5. Nitrogen-protected falling film concentration: Concentration temperature 37℃, vacuum degree 0.072MPa, nitrogen flow rate 20mL / min, concentrate concentration 8.5Brix, viscosity at 25℃ 16mPas, active ingredient loss rate 1.2%; S6. Pressure gradient non-thermal sterilization: low-pressure pre-sterilization 180MPa / 1 minute, medium-pressure stabilization 300MPa / 30 seconds, high-pressure inactivation 420MPa / 3 minutes, with a temperature of 1℃ throughout.

[0024] Example 2 S1. Antioxidant pretreatment: Matsutake alcohol content 0.17%, cap diameter 5.5cm, stipe length 11.5cm, diameter 2.6cm, time from harvesting to pretreatment 6.5 hours, washing water pressure 0.35MPa, washing time 40 seconds / time, drained moisture content 10%, slice thickness 3.5mm, ascorbate palmitate concentration in the compound pretreatment solution 0.02%, pH 6.6, material-to-liquid ratio 1:2.6g / mL, soaking temperature 3℃, time 15 minutes, fresh matsutake transportation temperature 2℃, buffer solution temperature difference from transportation temperature 1.5℃; S2. Vacuum-ultrasonic synergistic extraction: Spread thickness 1.5cm, vertical distance between vibrating plate and sheet 4cm, extraction temperature 4℃, vacuum degree 0.06MPa, ultrasonic frequency 30kHz, power density 0.35W / cm³. 2 The first 2 hours, 0.45 W / cm 2 For the next 2.0 hours, the heating rate was 0.75℃ / 10 minutes, the extraction time was 4.0 hours, the proportion of original solution collected was 72.5%, the start-stop time difference was 3.0 minutes, the nitrogen purging was 1.5 minutes / 30 minutes, the temporary storage temperature was 2℃, the pressure was 0.025MPa, the time was 2.5 hours, and the nitrogen flow rate was 7.5mL / min; S3, Gradient retention solid-liquid separation: Coarse filtration membrane operating pressure 0.20 MPa, flow rate 3.5 m / s² 3 / h, fine filtration membrane operating pressure 0.25MPa, flow rate 2.5m 3 / h; ceramic microfiltration membrane pore size 65nm, operating pressure 0.35MPa, flow rate 1.75m / h. 3 / h, filtrate temperature 3℃; S4. Modified membrane separation and purification: Ultrafiltration membrane pore size 15nm, molecular weight cutoff 20kDa, transmembrane pressure difference 0.065MPa, cross-flow velocity 1.25m / s, total content of three types of active ingredients in the filtrate 0.235%, protein rejection rate 98.0%; S5. Nitrogen-protected falling film concentration: Concentration temperature 39.5℃, vacuum degree 0.08MPa, nitrogen flow rate 25mL / min, concentrate concentration 10.0Brix, viscosity at 25℃ 20mPas, active ingredient loss rate 3.0%; S6. Pressure gradient non-thermal sterilization: low-pressure pre-sterilization 230MPa / 1 minute, medium-pressure stabilization 350MPa / 30 seconds, high-pressure inactivation 475MPa / 3 minutes, with a total temperature of 10℃.

[0025] Example 3 S1. Antioxidant pretreatment: Matsutake mushroom alcohol content 0.21%, cap diameter 6.5cm, stipe length 14cm, diameter 2.9cm, time from harvesting to pretreatment 9.5 hours, washing water pressure 0.4MPa, washing time 45 seconds / time, drained moisture content 14%, slice thickness 4.5mm, ascorbate palmitate concentration in the compound pretreatment solution 0.03%, pH 6.9, material-to-liquid ratio 1:3.1g / mL, soaking temperature 5℃, soaking time 18 minutes; fresh matsutake mushroom transportation temperature 4℃, buffer solution temperature difference from transportation temperature 2.5℃; S2. Vacuum-ultrasonic synergistic extraction: Spread thickness 2cm, vertical distance between vibrating plate and sheet 5cm, extraction temperature 7℃, vacuum degree 0.07MPa, ultrasonic frequency 40kHz, power density 0.4W / cm³. 2 First 2 hours, 0.5 W / cm 2 After 2.8 hours, the heating rate was 0.9℃ / 10 minutes, the extraction time was 4.8 hours, the original solution collection rate was 74%, the start-stop time difference was 4.5 minutes, the nitrogen purging was 2 minutes / 30 minutes, the temporary storage temperature was 3℃, the pressure was 0.028MPa, the time was 3.5 hours, and the nitrogen flow rate was 10mL / min. S3, Gradient retention solid-liquid separation: Coarse filtration membrane operating pressure 0.24 MPa, flow rate 3.9 m / s² 3 / h, fine filtration membrane operating pressure 0.29MPa, flow rate 2.9m 3 / h, ceramic microfiltration membrane pore size 80nm, operating pressure 0.4MPa, flow rate 2.0m 3 / h, filtrate temperature 5℃; S4. Modified membrane separation and purification: Ultrafiltration membrane pore size 19nm, molecular weight cutoff 29kDa; transmembrane pressure difference 0.09MPa, cross-flow velocity 1.5m / s, total content of three types of active ingredients in the filtrate 0.29%, protein rejection rate 98.8%; S5. Nitrogen-protected falling film concentration: Concentration temperature 42℃, vacuum degree 0.088MPa, nitrogen flow rate 30mL / min, concentrate concentration 11.5Brix, viscosity at 25℃ 24mPas, active ingredient loss rate 4.8%; S6. Pressure gradient non-thermal sterilization: low-pressure pre-sterilization 280MPa / 1 minute, medium-pressure stabilization 400MPa / 30 seconds, high-pressure inactivation 530MPa / 3 minutes, with a total temperature of 19℃.

[0026] Comparative Example 1 Unlike Example 2, ascorbate palmitate in the composite pretreatment solution was omitted, while the remaining process parameters were the same as in Example 2.

[0027] Comparative Example 2 Unlike Example 2, conventional ultrasonic extraction was used instead of vacuum ultrasonic synergistic extraction, while all other process parameters remained the same as in Example 2.

[0028] Comparative Example 3 Unlike Example 2, two-stage filtration replaces the gradient retention three-stage filtration, while the remaining process parameters are the same as in Example 2.

[0029] Comparative Example 4 Unlike Example 2, a conventional polyethersulfone film is used instead of the polyethersulfone and graphene composite modified film, while the other process parameters are the same as in Example 2.

[0030] Comparative Example 5 Unlike Example 2, conventional vacuum concentration replaces nitrogen-protected falling film concentration, while all other process parameters remain the same as in Example 2.

[0031] Comparative Example 6 Unlike Example 2, single pressure sterilization replaces pressure gradient non-thermal sterilization, while all other process parameters remain the same as in Example 2.

[0032] Table 1. Performance test data for Examples 1-3 unit Example 1 Example 2 Example 3 Total content of active ingredients % 0.180 0.235 0.290 Matsutake alcohol retention rate % 92.3 96.8 95.5 Protein retention rate % 97.2 98.0 98.8 Impurity removal rate % 99.4 99.7 99.6 Loss rate of active ingredients % 1.2 3.0 4.8 Concentrate viscosity (25℃) mPas 16.0 20.0 24.0 Sterilization pass rate % 98.5 99.8 99.5 Table 2, Performance test data for Comparative Examples 1-6 unit Comparative Example 1 Comparative Example 2 Comparative Example 3 Comparative Example 4 Comparative Example 5 Comparative Example 6 Total content of active ingredients % 0.201 0.176 0.228 0.214 0.209 0.217 Matsutake alcohol retention rate % 85.7 88.2 95.1 91.3 89.6 87.9 Protein retention rate % 97.8 97.5 97.6 92.5 97.9 97.7 Impurity removal rate % 99.6 99.5 98.2 99.6 99.6 99.6 Loss rate of active ingredients % 7.5 6.8 3.5 4.2 8.3 7.1 Concentrate viscosity mPas 19.8 19.5 20.2 20.1 20.3 20.2 Sterilization pass rate % 99.6 99.4 99.2 99.5 99.3 95.8 Based on the differences in Examples 1-3, Comparative Examples 1-6, and the performance test data table, it can be seen that the core advantage of this invention stems from the synergistic effect of six key processes: antioxidant protection of the composite pretreatment solution, vacuum ultrasonic synergistic extraction, gradient retention three-stage filtration, polyethersulfone and graphene composite modified membrane separation, nitrogen-protected falling film concentration, and pressure gradient non-thermal sterilization, as well as the precise adaptation of parameters in each step. These processes have a significant positive impact on the retention rate of active ingredients, extraction purity, impurity removal rate, stability, and sterilization safety, and achieve triple synergistic optimization of antioxidant and extraction efficiency, impurity classification and removal and separation accuracy, and inert protection and component stability.

[0033] Comparative Examples 1-6 all experienced a targeted decline in performance due to the absence or replacement of a single core process: removal of ascorbyl palmitate reduced matsutake alcohol retention to 85.7% and increased active ingredient loss to 7.5%; ordinary ultrasound replaced vacuum ultrasound, resulting in a total active ingredient content of only 0.176%; elimination of ceramic microfiltration membrane reduced impurity removal rate to 98.2%; ordinary polyethersulfone membrane resulted in a protein retention rate of only 92.5%; concentration without nitrogen protection resulted in an ingredient loss rate of 8.3%; and single pressure sterilization resulted in a matsutake alcohol retention rate of 87.9% and a sterilization pass rate of only 95.8%, fully demonstrating the indispensability of each process step.

[0034] Examples 1-3 all exhibited excellent performance: total active ingredient content of 0.18%-0.29%, matsutake alcohol retention rate of 92.3%-96.8%, active ingredient loss rate of 1.2%-4.8%, protein retention rate of 97.2%-98.8%, impurity removal rate of 99.4%-99.7%, and sterilization qualification rate of 98.5%-99.8%. Among them, Example 2 achieved the best performance balance, with a total active ingredient content of 0.235%, matsutake alcohol retention rate of 96.8%, and a loss rate of only 3.0%, fully meeting the standards for component stability and safety. Example 1 maintained the advantage of low loss rate with minimum parameters, while Example 3 achieved high component enrichment with maximum parameters. Together, they verified the rationality and application flexibility of the parameter range of the present invention.

[0035] The compatibility of parameters at each step is crucial for performance assurance: S1 pretreatment solution pH matches the osmotic pressure of matsutake cells; S2 ultrasonic parameters are adapted to the dissolution rate of active ingredients; S3 filtration membrane pore size gradient corresponds to impurity particle size distribution; S4 modified membrane parameters match the molecular weight of the target component; S5 concentration conditions match the thermal stability of the component; and S6 sterilization gradient is adapted to the structural tolerance of the component. Example 2 achieves an optimal balance of efficiency, purity, and safety through precise adaptation of parameters throughout the entire process. The synergistic process system of Examples 1-3 avoids efficiency or cost issues caused by parameter boundaries and covers the needs of different application scenarios.

[0036] In summary, this invention effectively solves the pain points of traditional extraction methods, such as large loss of active ingredients, low purity, incomplete removal of impurities, and poor stability, significantly improving the utilization efficiency of matsutake mushroom resources. Example 2 has the best cost performance and scalability potential, while the parameter flexibility of Examples 1-3 can be adapted to the needs of multiple scenarios such as deep processing, health product raw materials, and food additives, providing a replicable technical solution for the efficient and standardized extraction of active ingredients from matsutake mushrooms.

[0037] Table 3, Performance Indicators of Tables 1 and 2 (GB Testing Standards) GB testing standards Total content of active ingredients GB / T22244-2008, GB5009.5-2016, GB / T15672-2009 Matsutake alcohol retention rate Derived based on GB / T22244-2008 Protein retention rate GB5009.5-2016 Impurity removal rate GB5009.88-2014, GB5009.9-2016 Loss rate of active ingredients Derived based on the corresponding active ingredient detection standards Concentrate viscosity GB / T10247-2008 Sterilization pass rate GB4789.2-2016 Table 4. Test methods corresponding to GB testing standards Detection methods Total content of active ingredients Matsutake alcohol: High performance liquid chromatography, C18 column, methanol and water (70:30) as mobile phase, detection wavelength 220 nm; Peptides: Folin-Ciocalteu method, bovine serum albumin as standard, colorimetric determination at 750 nm; Polysaccharides: Anthrone and sulfuric acid method, glucose as standard, colorimetric determination at 620 nm; The sum is the total content. Matsutake alcohol retention rate Retention rate = (matsutake alcohol content after extraction / initial matsutake alcohol content in raw material) × 100%. Both raw material and extracted samples were determined according to GB / T22244-2008. Protein retention rate Retention rate = (1 - protein content in filtrate / protein content in extract) × 100%, protein content determined by Kjeldahl method. Impurity removal rate Dietary fiber: Enzymatic gravimetric method, weighing the residual amount after removing protein and starch; Starch: Determining the content difference before and after acid hydrolysis; Colloid: Centrifugation method (8000 r / min, 20 min), weighing the precipitate; Total removal rate = (1 - total impurities in filtrate / total impurities in original solution) × 100% Loss rate of active ingredients Loss rate = (Total active ingredient content before each process stage - Total active ingredient content after each process stage) / Total active ingredient content before each process stage × 100%, with a focus on the concentration and sterilization stages. Concentrate viscosity For the rotational viscometer method, select a suitable rotor, rotate at 60 r / min, and take the reading after stabilizing at a constant temperature of 25℃ for 30 seconds. Sterilization pass rate <![CDATA[Plate counting method. Take 10 g of the sample and dilute it to 10 -1 -10 -3 concentration, pour nutrient agar medium, incubate at 36 °C for 48 h, count the number of colonies, and the proportion of samples meeting the standard is the pass rate]]> Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. A low-temperature, high-efficiency extraction method for active ingredients from matsutake mushrooms, characterized in that, Includes the following steps: S1. Antioxidant pretreatment: Select qualified fresh matsutake mushrooms, wash them with reverse osmosis purified water, drain them in a sterile environment, and then slice them to obtain matsutake slices. Place the matsutake slices in a sterile buffer solution containing antioxidants and soak them at low temperature to maintain the stability of the matsutake cell structure. S2, Vacuum-ultrasonic synergistic extraction: The matsutake slices soaked in S1 are transferred to a vacuum ultrasonic extraction device. Under low temperature and vacuum conditions, combined with ultrasonic assistance, drying and active ingredient extraction are carried out simultaneously, and the matsutake small molecular cluster cell original solution is collected in real time. S3. Gradient retention solid-liquid separation: First, a large-pore coarse filtration membrane is used to remove large particles of dietary fiber, then a fine filtration membrane is used to retain starch particles, and finally a ceramic microfiltration membrane is used to retain colloidal impurities. The three-stage filtration process yields a highly clear filtrate. S4. Modified membrane separation and purification: The filtrate from the three-stage filtration in S3 is passed into a surface-modified ultrafiltration membrane system to selectively retain large protein molecules while simultaneously retaining small molecule active ingredients, and the filtrate rich in matsutake alcohol, peptides and polysaccharides is collected. S5. Nitrogen-protected falling film concentration: The filtrate from S4 is concentrated under low temperature and vacuum conditions, with nitrogen gas introduced throughout the process to isolate oxygen and prevent the active ingredients from oxidizing and degrading, resulting in a highly active matsutake mushroom concentrate. S6. Pressure gradient non-thermal sterilization: The concentrated stock solution of S5 is treated with a three-stage pressure gradient to maximize the retention of active ingredients while ensuring the sterilization effect, so as to obtain high-purity matsutake mushroom extract.

2. The low-temperature, high-efficiency extraction method for active ingredients of matsutake mushroom according to claim 1, characterized in that: In S1, the thickness of the matsutake slices is 2.5-4.5 mm; The qualified fresh matsutake mushrooms contain 0.14% to 0.21% matsutake alcohol, the caps are not fully open and the diameter is 4.5 to 6.5 cm, the stem length is 9 to 14 cm and the diameter is 2.3 to 2.9 cm, and the total time from picking to S1 pretreatment is 3.5 to 9.5 hours. The antioxidant is 0.01-0.03% ascorbate palmitate, which is mixed with phosphate buffer to form a composite pretreatment solution; The pH of the composite pretreatment solution was 6.4-6.9, the mass-to-volume ratio of the solution to the matsutake slices was 1:2.11:3.1 g / mL, the soaking temperature was 15℃, the soaking time was 12-18 minutes, and the difference between the temperature of the buffer solution and the transport temperature of the fresh matsutake was 0.5-2.5℃.

3. The low-temperature, high-efficiency extraction method for active ingredients of matsutake mushroom according to claim 1, characterized in that: In step S1, the reverse osmosis purified water cleaning pressure is 0.3-0.4 MPa, the cleaning is performed twice, and the cleaning time for each cleaning is 35-45 seconds. After draining, the surface moisture content of the matsutake mushroom slices was 6%-14%.

4. The low-temperature, high-efficiency extraction method for active ingredients of matsutake mushroom according to claim 1, characterized in that: In S2, the low temperature condition is 17°C, and the vacuum condition is a vacuum degree of 0.05-0.07 MPa; Ultrasound-assisted parameters include: frequency 2040kHz, power density 0.3-0.5W / cm². 2 The ultrasound is activated intermittently. The heating rate of the vacuum extraction equipment is 0.6-0.9℃ / 10 minutes, the total drying and extraction time is 3.2-4.8 hours, the original liquid collected simultaneously accounts for 71%-74% of the total cell liquid content of matsutake mushrooms, and the start-stop time difference between the original liquid collection and the drying and extraction is 1.5-4.5 minutes, which increases the dissolution rate of active ingredients by 15%-20%.

5. The low-temperature, high-efficiency extraction method for active ingredients of matsutake mushroom according to claim 1, characterized in that: In step S2, the collected stock solution is temporarily stored in a sterile, sealed container under the following conditions: Temperature 13℃, pressure 0.022-0.028MPa, temporary storage time 1.5-3.5 hours, immediately proceed to S3 gradient retention filtration step; A small amount of nitrogen gas was continuously introduced during the temporary storage process at a flow rate of 510 mL / min.

6. The low-temperature, high-efficiency extraction method for active ingredients of matsutake mushroom according to claim 1, characterized in that: In step S3, the gradient filtration includes: a coarse filter membrane with a pore size of 0.5 μm, an operating pressure of 0.16-0.24 MPa, and a flow rate of 3.1-3.9 m / s². 3 / h; The fine filtration membrane has a pore size of 0.2 μm, an operating pressure of 0.21-0.29 MPa, and a flow rate of 2.1-2.9 m / s. 3 / h; The ceramic microfiltration membrane has a pore size of 50-80 nm, an operating pressure of 0.3-0.4 MPa, and a flow rate of 1.5-2.0 m / s. 3 / h; During the three-stage filtration process, the filtrate temperature is maintained at 15℃, which is consistent with the collection temperature of the S2 stock solution, and the impurity removal rate is 99.2%.

7. The low-temperature, high-efficiency extraction method for active ingredients of matsutake mushroom according to claim 1, characterized in that: In S4, the surface-modified ultrafiltration membrane is a polyethersulfone and graphene composite modified membrane with a pore size of 1119 nm and a molecular weight cutoff of 1129 kDa for protein impurities. The transmembrane pressure difference of the ultrafiltration membrane system was controlled at 0.04-0.09 MPa. Cross-flow filtration was adopted with a cross-flow velocity of 11.5 m / s. The total content of matsutake alcohol, matsutake polypeptide and matsutake polysaccharide in the filtrate was 0.18%-0.29%, the protein retention rate was 97.2%-98.8%, and the permeation rate of active ingredients was increased by 12%-18%.

8. The low-temperature, high-efficiency extraction method for active ingredients of matsutake mushroom according to claim 1, characterized in that: In step S5, the temperature of vacuum falling film concentration is 3742℃ and the vacuum degree is 0.072-0.088MPa; The nitrogen protection flow rate is 2030 mL / min, covering the surface of the concentrate to form an inert atmosphere; The concentration of the concentrated matsutake mushroom concentrate was 8.5-11.5 Brix, the loss rate of active ingredients during the concentration process was 1.2%-4.8%, and the viscosity at 25℃ was 1624 mPas.

9. The low-temperature, high-efficiency extraction method for active ingredients of matsutake mushroom according to claim 1, characterized in that: In S6, the three-stage pressure gradient consists of three phases: low-pressure pre-sterilization, medium-pressure stabilization, and high-pressure inactivation. Low-pressure pre-sterilization: 180-280 MPa, processing time: 1 minute; Medium-pressure stable pressure 300-400MPa, processing time 30 seconds; High-pressure inactivation: 420-530 MPa, holding time: 3 minutes; The entire process is carried out at a temperature of 119℃, using a step-by-step pressurization method.

10. The low-temperature, high-efficiency extraction method for active ingredients of matsutake mushroom according to claim 1, characterized in that: In S2, vacuum ultrasonic synergistic extraction also includes: the matsutake mushroom slices are spread to a thickness of 12cm in the vacuum ultrasonic extraction equipment; Nitrogen purging was performed every 30 minutes during the extraction process, with a purging time of 12 minutes. After purging, a vacuum of 0.05-0.07 MPa was maintained, and the ultrasonic power was dynamically adjusted according to the extraction time, maintaining a power density of 0.3-0.4 W / cm³ for the first 2 hours. 2 After 1.2 to 2.8 hours, the concentration was adjusted to 0.4 to 0.5 W / cm². 2 .

Images