A method for extracting a high-activity ginseng extract

Abstract

The present application belongs to the technical field of traditional Chinese medicine extraction, and particularly relates to an extraction method of high-activity ginseng extract, which is used for improving the conversion and extraction efficiency of rare ginsenosides Rg1, Rg3, Rg5, Rh2 and Compound K. The extraction method combines microbial fermentation, electrostatic field wall breaking and supercritical CO2 extraction. The microbial fermentation treatment is mainly used for promoting the synthesis of ginsenosides, the high-voltage pulse electric field induces the electroporation effect of ginseng cells to increase the permeability of cell membrane and cell wall, the supercritical CO2 extraction has high cell permeability to improve the extraction rate and purity of ginsenosides, and in addition, the catalytic aid added in the extraction process is helpful to the conversion and catalysis of rare ginsenoside components.

Description

Technical Field

[0001] This invention belongs to the field of traditional Chinese medicine extraction technology, specifically relating to a method for extracting highly active ginseng extract. Background Technology

[0002] Research on the extraction process of ginsenosides is a key area in the development of active ingredients in traditional Chinese medicine and modern formulation technology. Traditional extraction methods, such as decoction, maceration, and reflux extraction, while simple in equipment and convenient in operation, generally have low extraction efficiency, often requiring large amounts of solvent and time. Furthermore, continuous heating can easily cause hydrolysis or isomerization of heat-sensitive saponins, leading to a decrease in the activity of the target product. In recent years, modern extraction technologies, such as ultrasonic extraction, microwave-assisted extraction, and supercritical CO2 fluid extraction, have shown significant progress in improving extraction rates and reducing solvent consumption. For example, ultrasonic extraction promotes cell rupture through cavitation, microwaves accelerate component dissolution through internal heating, and supercritical CO2, with its high permeability and low-temperature operation advantages, helps maintain component stability. However, these technologies still face common challenges in practical applications: on the one hand, their extraction processes are mostly based on physical principles and lack selectivity for saponin components, making it difficult to selectively enrich rare ginsenosides; on the other hand, since the content of these rare saponins in the raw materials is extremely low and they are mostly tightly bound to the cell structure, it is difficult to achieve their efficient release and transformation by physical means alone, which often leads to the difficulty in significantly increasing the proportion of rare saponins in the final extract, thus restricting the development of related high value-added products. Summary of the Invention

[0003] In view of the above situation, the present invention provides an extraction method for highly active ginseng extract, aiming to construct a green and efficient ginseng extraction process to improve the content and extraction rate of highly active rare ginsenosides in ginseng extract. By combining microbial fermentation, electrostatic field cell disruption and supercritical CO2 extraction, the efficient dissolution and activity stability of ginsenosides under mild conditions are achieved.

[0004] To achieve the above objectives, the technical solution adopted by the present invention is as follows:

[0005] This invention provides a method for extracting highly active ginseng extract, specifically including the following steps:

[0006] Step 1: Take ginseng, wash, dry, and crush it to obtain ginseng powder. Use a high-yield strain of β-glucosidase as the fermentation strain. After cultivation and amplification, obtain seed liquid. Moisten the ginseng powder with purified water and then perform high-pressure steam sterilization to avoid contamination by other microorganisms. Obtain sterilized material. Spray the seed liquid onto the surface of the sterilized material and place it in a fermentation chamber for 5 days to promote the production of rare saponin components Rg1, Rg3, Rh2, and Rg5 in ginseng. Dry at 90℃ to inactivate microorganisms and enzymes to obtain fermented powder.

[0007] Step 2: Spread the fermentation powder evenly in a parallel plate electrode device, and treat it with a high-voltage pulsed electric field: field strength 20-25kV / cm, pulse frequency 1000 Hz, time 10-15 min, temperature 40℃. The high-voltage pulsed electric field induces electroporation in ginseng cells, increases the permeability of cell membrane and cell wall, promotes the diffusion of intracellular contents to the extracellular medium, and makes polar saponin components easier to dissolve, thus obtaining pretreated fermentation powder.

[0008] Step 3: Extract the pretreated fermentation powder using a catalytic aid and supercritical CO2 extraction. Mix the pretreated fermentation powder, catalytic aid, and anhydrous ethanol in a mass ratio of 10:1.5:1 and add them to the high-pressure extraction vessel of the supercritical extraction device. The solubilizing and inclusion effects provided by the catalytic aid are conducive to promoting the dissolution of saponin components. The added glycosidase can promote the directional catalytic conversion of native saponin substrates Rb1, Rc, and Rd into rare ginsenosides Rg3 and Compound K. Use CO2 as the extractant, set the extraction pressure to 25-35 MPa, CO2 flow rate to 150 L / h, extraction temperature to 45-55℃, and extraction time to 1-2 h. Under supercritical conditions, CO2 fluid has both high diffusivity and strong dissolving ability, which can efficiently penetrate into the interior of the material and selectively extract the target saponin components. After extraction, the supercritical CO2 fluid containing ginseng extract enters the separation vessel, and the CO2 is vaporized and separated by depressurization to collect the extract.

[0009] Step 4: The extract was ultrafiltered and centrifuged using an ultrafiltration tube with a molecular weight of 10 kDa to remove large molecular proteins, colloids and possible microbial residues, and the resulting pre-filtrate was loaded onto a macroporous adsorption resin column for purification. The pre-filtrate was first eluted with purified water to remove residual salts, sugars and other water-soluble impurities and some water-soluble catalyst components, and then eluted with 70% ethanol solution. The eluent was collected.

[0010] Step 5: Pass the eluent through a decolorizing resin column to remove impurities such as pigments and phenols, and obtain a purified solution. Concentrate and dry the purified solution to obtain a highly active ginseng extract.

[0011] Furthermore, the catalyst comprises the following raw materials by weight percentage: 1% polysorbate 80, 1.5% β-cyclodextrin, 0.4% citric acid, 0.3% sodium citrate, 0.4% calcium chloride, 1% β-glucosidase, 1% β-xylosidase, 4.4% microcrystalline cellulose and 90% purified water.

[0012] Furthermore, the high-yielding β-glucosidase strain is selected from any one of Lactobacillus plantarum LP115, Lactobacillus plantarum subsp. plantarum ATCC14917, and Lactobacillus fermentum CECT5716.

[0013] The beneficial effects achieved by this invention are as follows:

[0014] The extraction method for highly active ginseng extract provided by this invention combines microbial fermentation transformation, high-voltage pulsed electric field cell disruption with auxiliary agents, and gas-assisted extraction to achieve efficient, targeted, and green preparation of ginseng active ingredients, especially rare ginsenosides. Microorganisms, under mild conditions, biocatalyze the conversion of abundant proto-ginsenosides in ginseng into highly active rare ginsenosides Rg1, Rg3, Rh2, and Rg5, enhancing the value and content of the target product. Furthermore, the electroporation effect generated by the high-voltage pulsed electric field deeply disrupts the ultrastructure of the cell walls and cell membranes of the fermented ginseng powder, significantly reducing the mass transfer resistance in subsequent extraction and creating conditions for the full dissolution of active ingredients. During the extraction process, the added auxiliary agents have multiple functions, including solubilization, inclusion, and catalytic transformation. β-glucosidase and β-xylosidase can cleave the glucose groups of the native saponin substrates Rb1, Rc, and Rd, promoting their targeted catalytic conversion into rare ginsenosides Rg3 and Compound. K enables a one-step extraction process combining enzymatic directional conversion and simultaneous solvent extraction. Supercritical CO2 fluid, with its extremely low viscosity and high permeability, rapidly penetrates the highly porous plant cell matrix, pretreated by microbial fermentation and an electric field. This facilitates the rapid and complete dissolution of saponins, particularly rare ginsenosides with moderate molecular weight and relatively low polarity, significantly improving extraction efficiency and yield. Simultaneously, the extraction process is conducted in a relatively mild and inert environment, effectively preventing the decomposition, isomerization, or oxidative inactivation of heat-sensitive saponins at high temperatures. This preserves their natural conformation and bioactivity, greatly enhancing extraction efficiency and selectivity, resulting in ginseng extracts with light color, high purity, and a significantly increased proportion of rare saponins. Attached Figure Description

[0015] Figure 1 This is a process flow diagram of the extraction method for the highly active ginseng extract of the present invention;

[0016] Figure 2 The results are ultraviolet full-wavelength scans of the ginsenoside reference standard and the highly active ginseng extract extracted in Example 4.

[0017] Figure 3 The standard curve for ginsenoside reference standards;

[0018] Figure 4 The results of total saponin content and total saponin extraction rate of the highly active ginseng extracts extracted in Examples 2-6 are as follows;

[0019] Figure 5 The results show the component analysis of the ginseng extracts extracted in Comparative Example 1 and Example 4. Detailed Implementation

[0020] The technical solutions of the present invention will be clearly and completely described below with reference to the accompanying drawings of the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without creative effort are within the scope of protection of the present invention.

[0021] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as those familiar to those skilled in the art. Furthermore, any methods and materials similar to or equivalent to those described herein may be applied to this invention. The preferred embodiments and materials described herein are for illustrative purposes only and do not limit the scope of this application.

[0022] Unless otherwise specified, the methods described in the following embodiments are conventional; the process flow diagram of the extraction method of a highly active ginseng extract of the present invention is as follows. Figure 1 As shown; unless otherwise specified, all materials used in the following examples are new materials purchased from the market. All parts mentioned in the following examples are by weight. Specifically, all medicinal parts of ginseng used are processed Chinese medicinal slices; the β-glucosidase used has an enzyme activity of 20000 U / g; the β-xylosidase used has an enzyme activity of 18000 U / g; and the *Lactobacillus plantarum* LP115, *Lactobacillus plantarum* subsp. *plantarum* ATCC14917, and *Lactobacillus fermentum* CECT5716 used are all commercially available strains.

[0023] Example 1: This example provides a catalytic aid comprising the following raw materials in parts by weight: 1 part polysorbate 80, 1.5 parts β-cyclodextrin, 0.4 parts citric acid, 0.3 parts sodium citrate, 0.4 parts calcium chloride, 1 part β-glucosidase, 1 part β-xylosidase, 4.4 parts microcrystalline cellulose, and 90 parts purified water.

[0024] Example 2: This example provides a method for extracting highly active ginseng extract, specifically including the following steps:

[0025] Step 1: Take 214 portions of ginseng body, wash, dry, and pulverize them. Pass them through a 60-mesh sieve to obtain ginseng powder. Use Lactobacillus plantarum LP115 as the fermentation inoculum. After activation, culture, and amplification, obtain seed liquid. The viable count of the seed liquid is approximately 1×10⁻⁶. 8CFU / mL, after moistening the ginseng powder with purified water, it was sterilized by high-pressure steam at 121℃ and 0.1 MPa for 30 min to obtain sterilized material. The seed liquid was sprayed onto the surface of the sterilized material at a mass-volume ratio of ginseng powder: 100 g: 4 mL, and then placed in a fermentation chamber for static fermentation at 37℃ and 85% RH for 5 days, turning the pile once every 24 h during the period. After the fermentation was completed, it was dried at 90℃ to a moisture content of 8.4% to obtain fermented powder.

[0026] Step 2: Spread the yeast powder evenly in the parallel plate electrode device with a thickness of about 2 mm, and treat it with a high-voltage pulse electric field: field strength 20 kV / cm, pulse frequency 1000 Hz, time 15 min, temperature 40℃ to obtain pretreated yeast powder.

[0027] Step 3: Take 100 parts of pretreated fermentation powder, 15 parts of catalyst and 10 parts of anhydrous ethanol respectively, mix them and add them to the high pressure extraction vessel of the supercritical extraction device for supercritical CO2 extraction. Set the extraction pressure to 25 MPa, CO2 flow rate to 150 L / h, extraction temperature to 50℃, and extraction time to 2 h. After extraction, reduce the pressure to 5 MPa to vaporize and separate the CO2, and collect the extract. The extract is rich in ginsenosides.

[0028] Step 4: The extract was ultrafiltered and centrifuged for 20 min at 4℃ and 8000 rpm using an ultrafiltration tube with a molecular weight of 10 kDa to obtain the initial filtrate. The initial filtrate was loaded into a pretreated D101 macroporous adsorption resin column (height-to-diameter ratio 5:1) at a flow rate of 2 BV / h for purification. First, it was eluted with 5 BV of purified water at the same flow rate, and then eluted with 4 BV of 70% ethanol solution at a flow rate of 1 BV / h. The eluent was collected.

[0029] Step 5: Pass the eluent through an ADS-7 decolorizing resin column at a flow rate of 2 BV / h, collect the purified solution, concentrate the purified solution and spray dry it to obtain a highly active ginseng extract, weighing 12.2 g (12.2 parts).

[0030] Example 3: This example provides a method for extracting highly active ginseng extract, specifically including the following steps:

[0031] Step 1: Take 155 parts of ginseng rootlets, wash, dry, and pulverize them. Pass the powder through a 60-mesh sieve to obtain ginseng powder. Use Lactobacillus plantarum subsp. plantarum ATCC14917 as the fermentation strain. After activation, culture, and amplification, obtain seed liquid. The viable count of the seed liquid is approximately 1×10⁻⁶. 8CFU / mL, after moistening the ginseng powder with purified water, it was sterilized by high-pressure steam at 121℃ and 0.1 MPa for 30 min to obtain sterilized material. The seed liquid was sprayed onto the surface of the sterilized material at a mass-volume ratio of ginseng powder: 100 g: 4 mL, and then placed in a fermentation chamber for static fermentation at 37℃ and 85% RH for 5 days, turning the pile once every 24 h. After fermentation, it was dried at 90℃ to a moisture content of 6.9% to obtain fermented powder.

[0032] Step 2: Spread the yeast powder evenly in the parallel plate electrode device with a thickness of about 2 mm, and treat it with a high-voltage pulse electric field: field strength 20 kV / cm, pulse frequency 1000 Hz, time 15 min, temperature 40℃ to obtain pretreated yeast powder.

[0033] Step 3: Take 100 parts of pretreated fermentation powder, 15 parts of catalyst and 10 parts of anhydrous ethanol respectively, mix them and add them to the high-pressure extraction vessel of the supercritical extraction device for supercritical CO2 extraction. Set the extraction pressure to 25 MPa, CO2 flow rate to 150 L / h, extraction temperature to 45℃, and extraction time to 2 h. After extraction, reduce the pressure to 5 MPa to vaporize and separate the CO2, and collect the extract.

[0034] Step 4: The extract was ultrafiltered and centrifuged for 20 min at 4℃ and 8000 rpm using an ultrafiltration tube with a molecular weight of 10 kDa to obtain the initial filtrate. The initial filtrate was loaded into a pretreated D101 macroporous adsorption resin column (height-to-diameter ratio 5:1) at a flow rate of 2 BV / h for purification. First, it was eluted with 5 BV of purified water at the same flow rate, and then eluted with 4 BV of 70% ethanol solution at a flow rate of 1 BV / h. The eluent was collected.

[0035] Step 5: Pass the eluent through an ADS-7 decolorizing resin column at a flow rate of 2 BV / h, collect the purified solution, concentrate the purified solution and spray dry it to obtain a highly active ginseng extract, weighing 9.7 g (9.7 parts).

[0036] Example 4: This example provides a method for extracting highly active ginseng extract, specifically including the following steps:

[0037] Step 1: Take 161 portions of ginseng rootlets, wash, dry, and pulverize them. Pass the powder through a 60-mesh sieve to obtain ginseng powder. Use Lactobacillus plantarum LP115 as the fermentation starter. After activation, culture, and amplification, obtain seed liquid. The viable count of the seed liquid is approximately 1×10⁻⁶. 8CFU / mL, after moistening the ginseng powder with purified water, it was sterilized by high-pressure steam at 121℃ and 0.1 MPa for 30 min to obtain sterilized material. The seed liquid was sprayed onto the surface of the sterilized material at a mass-volume ratio of ginseng powder: 100 g: 4 mL, and then placed in a fermentation chamber for static fermentation at 37℃ and 85% RH for 5 days, turning the pile once every 24 h. After fermentation, it was dried at 90℃ to a moisture content of 7.4% to obtain fermented powder.

[0038] Step 2: Spread the yeast powder evenly in the parallel plate electrode device with a thickness of about 2 mm, and treat it with a high-voltage pulse electric field: field strength 25 kV / cm, pulse frequency 1000 Hz, time 10 min, temperature 40℃, to obtain pretreated yeast powder.

[0039] Step 3: Take 100 parts of pretreated fermentation powder, 15 parts of catalyst and 10 parts of anhydrous ethanol respectively, mix them and add them to the high-pressure extraction vessel of the supercritical extraction device for supercritical CO2 extraction. Set the extraction pressure to 30 MPa, CO2 flow rate to 150 L / h, extraction temperature to 50℃, and extraction time to 1.5 h. After extraction, reduce the pressure to 5 MPa to vaporize and separate the CO2, and collect the extract.

[0040] Step 4: The extract was ultrafiltered and centrifuged for 20 min at 4℃ and 8000 rpm using an ultrafiltration tube with a molecular weight of 10 kDa to obtain the initial filtrate. The initial filtrate was loaded into a pretreated D101 macroporous adsorption resin column (height-to-diameter ratio 5:1) at a flow rate of 2 BV / h for purification. First, it was eluted with 5 BV of purified water at the same flow rate, and then eluted with 4 BV of 70% ethanol solution at a flow rate of 1 BV / h. The eluent was collected.

[0041] Step 5: Pass the eluent through an ADS-7 decolorizing resin column at a flow rate of 2 BV / h, collect the purified solution, concentrate the purified solution and spray dry it to obtain a highly active ginseng extract, weighing 22.85 g (22.85 parts).

[0042] Example 5: This example provides a method for extracting highly active ginseng extract, specifically including the following steps:

[0043] Step 1: Take 143 portions of ginseng rootlets, wash, dry, and pulverize them. Pass the powder through a 60-mesh sieve to obtain ginseng powder. Use *Lactobacillus mucilaginosus* CECT5716 as the fermentation strain. After activation, culture, and amplification, obtain seed liquid. The viable count of the seed liquid is approximately 1 × 10⁻⁶. 8CFU / mL, after moistening the ginseng powder with purified water, it was sterilized by high-pressure steam at 121℃ and 0.1 MPa for 30 min to obtain sterilized material. The seed liquid was sprayed onto the surface of the sterilized material at a mass-volume ratio of ginseng powder: 100 g: 4 mL, and then placed in a fermentation chamber for static fermentation at 37℃ and 85% RH for 5 days, turning the pile once every 24 h. After fermentation, it was dried at 90℃ to a moisture content of 6.5% to obtain fermented powder.

[0044] Step 2: Spread the yeast powder evenly in the parallel plate electrode device with a thickness of about 2 mm, and treat it with a high-voltage pulse electric field: field strength 22 kV / cm, pulse frequency 1000 Hz, time 12 min, temperature 40℃ to obtain pretreated yeast powder.

[0045] Step 3: Take 100 parts of pretreated fermentation powder, 15 parts of catalyst and 10 parts of anhydrous ethanol respectively, mix them and add them to the high pressure extraction vessel of the supercritical extraction device for supercritical CO2 extraction. Set the extraction pressure to 35 MPa, CO2 flow rate to 150 L / h, extraction temperature to 55℃, and extraction time to 1 h. After extraction, reduce the pressure to 5 MPa to vaporize and separate the CO2, and collect the extract.

[0046] Step 4: The extract was ultrafiltered and centrifuged for 20 min at 4℃ and 8000 rpm using an ultrafiltration tube with a molecular weight of 10 kDa to obtain the initial filtrate. The initial filtrate was loaded into a pretreated D101 macroporous adsorption resin column (height-to-diameter ratio 5:1) at a flow rate of 2 BV / h for purification. First, it was eluted with 5 BV of purified water at the same flow rate, and then eluted with 4 BV of 70% ethanol solution at a flow rate of 1 BV / h. The eluent was collected.

[0047] Step 5: Pass the eluent through an ADS-7 decolorizing resin column at a flow rate of 2 BV / h, collect the purified solution, concentrate the purified solution and spray dry it to obtain a highly active ginseng extract, weighing 14.54 g (14.54 parts).

[0048] Example 6: This example provides a method for extracting highly active ginseng extract, specifically including the following steps:

[0049] Step 1: Take 216 portions of whole ginseng, wash, dry, and pulverize them. Pass the powder through a 60-mesh sieve to obtain ginseng powder. Use Lactobacillus plantarum LP115 as the fermentation starter. After activation, culture, and amplification, obtain seed liquid. The viable count of the seed liquid is approximately 1×10⁻⁶. 8CFU / mL, after moistening the ginseng powder with purified water, it was sterilized by high-pressure steam at 121℃ and 0.1 MPa for 30 min to obtain sterilized material. The seed liquid was sprayed onto the surface of the sterilized material at a mass-volume ratio of ginseng powder: 100 g: 4 mL, and then placed in a fermentation chamber for static fermentation at 37℃ and 85% RH for 5 days, turning the pile once every 24 h. After fermentation, it was dried at 90℃ to a moisture content of 9.2% to obtain fermented powder.

[0050] Step 2: Spread the yeast powder evenly in the parallel plate electrode device with a thickness of about 2 mm, and treat it with a high-voltage pulse electric field: field strength 25 kV / cm, pulse frequency 1000 Hz, time 10 min, temperature 40℃, to obtain pretreated yeast powder.

[0051] Step 3: Take 100 parts of pretreated fermentation powder, 15 parts of catalyst and 10 parts of anhydrous ethanol respectively, mix them and add them to the high pressure extraction vessel of the supercritical extraction device for supercritical CO2 extraction. Set the extraction pressure to 30 MPa, CO2 flow rate to 150 L / h, extraction temperature to 55℃, and extraction time to 2 h. After extraction, reduce the pressure to 5 MPa to vaporize and separate the CO2, and collect the extract.

[0052] Step 4: The extract was ultrafiltered and centrifuged for 20 min at 4℃ and 8000 rpm using an ultrafiltration tube with a molecular weight of 10 kDa to obtain the initial filtrate. The initial filtrate was loaded into a pretreated D101 macroporous adsorption resin column (height-to-diameter ratio 5:1) at a flow rate of 2 BV / h for purification. First, it was eluted with 5 BV of purified water at the same flow rate, and then eluted with 4 BV of 70% ethanol solution at a flow rate of 1 BV / h. The eluent was collected.

[0053] Step 5: Pass the eluent through an ADS-7 decolorizing resin column at a flow rate of 2 BV / h, collect the purified solution, concentrate the purified solution and spray dry it to obtain a highly active ginseng extract, weighing 7.76 g (7.76 parts).

[0054] Comparative Example 1: The difference from Example 4 is that the medicinal materials were directly extracted by supercritical CO2 after being pulverized, without microbial fermentation, high-voltage electric field treatment, or catalytic agent treatment.

[0055] Total saponin content determination: The total saponin content of ginseng extract was determined using a U-3900 UV-Vis spectrophotometer. According to the standard for the determination of total ginseng saponin content, the reference standard was ginsenoside Re standard, with a purity of 96.9%, purchased from the China National Institutes for Food and Drug Control. The test samples were highly active ginseng extracts extracted in Examples 2-6, numbered 1-5 respectively. Solutions of the reference standard and test sample 3 (Example 4) were prepared at 1 mg / mL, and UV full-wavelength scanning was performed. The absorption spectra are shown below. Figure 2 .

[0056] Figure 2 The results showed that the absorption spectra of the reference standard and the test sample were basically the same, and both had maximum absorption at 560 nm. Therefore, 560 nm was selected as the detection wavelength for the total saponin content.

[0057] Preparation of the standard curve: Accurately weigh 10.14 mg of the reference standard and place it in a 10 mL volumetric flask. Dissolve it in methanol and dilute to the mark to prepare the reference standard stock solution. Accurately measure 20, 40, 80, 120, 160, and 200 μL of the above reference standard stock solution into stoppered test tubes. Evaporate the solvent at 60 °C. Accurately add 200 μL of 5% vanillin-glacial acetic acid solution and 800 μL of perchloric acid, shake well, incubate in a 60 °C water bath for 10 min, cool in an ice bath for 2 min, accurately add 4 mL of glacial acetic acid, shake well, and measure the absorbance at 560 nm. Plot the standard curve with concentration on the x-axis and absorbance on the y-axis. The results are shown in the figure. Figure 3 .

[0058] Determination of total saponin content in test samples: Take 10 mg of each of test samples 1-5 (Examples 2-6) into a 5 mL volumetric flask, add methanol and sonicate for 20 min, let stand at room temperature, then add methanol to the mark, shake well, filter through a 0.22 μm filter membrane, and take 50 μL to determine the absorbance using the same detection method as above. Substitute the values ​​into the standard curve to calculate the concentration, and calculate the total saponin content (%) in the high-activity ginseng extract = detection concentration (mg / mL) × 10 (dilution factor) × 5 mL / 10 mg × 100%, and the total saponin extraction rate (%) = ginseng extract mass × total saponin content / pretreated fermentation powder mass × 100%. The results are shown in the figure. Figure 4 .

[0059] Ginsenoside component analysis: The content of each saponin component in the highly active ginseng extracts obtained in Comparative Example 1 and Example 4 was analyzed by HPLC high-performance liquid chromatography. The HPLC chromatographic conditions are as follows: C 18The chromatographic column (4.6 mm × 250 mm, 5 μm), flow rate 1 mL / min, column temperature 30℃, injection volume 20 μL, detection wavelength 203 nm, mobile phase: acetonitrile (A) - 0.1% phosphoric acid water (B), gradient elution program: 1-10 min (20%A), 10-20 min (20%A→35%A), 20-60 min (35%A→60%A), 60-70 min (60%A), 70-80 min (60%A→90%A), 80-90 min (90%), 90-91 min (90%A→20%A), 91-100 min (20%A). The HPLC results are shown below. Figure 5 The results of the detection of some components of ginsenosides are shown in Table 1.

[0060] Table 1. Results of Ginseng Extract Content Determination

[0061]

[0062] Figure 2 The results showed that the maximum absorption wavelength of the reference standard and the test sample was at 560 nm, and their absorbance was similar.

[0063] Figure 3 The results show that the standard curve equation is A = 0.02281x - 0.03185, R0 2 =0.9994, indicating good linearity, suitable for content detection of test samples.

[0064] Figure 4 The results showed that the total ginsenoside content of the ginseng extracted in Examples 2-6 was greater than 85%, and the extraction efficiency was high. Among them, the total ginsenoside content and total ginsenoside extraction rate of the ginseng extract extracted in Example 4 were the highest, reaching 94% and 23%, respectively.

[0065] Figure 5 The results in Table 1 show that the content of each saponin component extracted in Example 4 was significantly increased compared with that in Comparative Example 1, indicating the specificity of the extraction process for saponin components. At the same time, the content of rare ginsenosides Rg1, Rg3, Rg5, Rh2, and Compound K increased, indicating that the microbial fermentation and catalytic aids had good catalytic efficiency for the conversion of rare ginsenosides.

[0066] 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.

[0067] The present invention and its embodiments have been described above. This description is not restrictive, and the accompanying drawings are only one embodiment of the present invention. The actual application is not limited to this. In conclusion, if those skilled in the art are inspired by this description and design similar methods and embodiments without departing from the spirit of the present invention, they should all fall within the protection scope of the present invention.

Claims

1. A method for extracting a highly active ginseng extract, characterized in that, Specifically, the following steps are included: Step 1: Take ginseng, wash it, dry it, and grind it into powder to obtain ginseng powder. Then, ferment the ginseng powder to obtain baking powder. Step 2: Treat the yeast powder with a high-voltage pulsed electric field to obtain pretreated yeast powder; Step 3: Mix the pretreated yeast, catalyst and anhydrous ethanol, and perform supercritical CO2 extraction to collect the extract; Step 4: The extract is subjected to ultrafiltration centrifugation, adsorption resin purification, and elution, and the eluent is collected. Step 5: Decolorize, concentrate, and dry the eluent to obtain a highly active ginseng extract; In step 1, the fermentation process uses a high-yield strain of β-glucosidase; The high-yielding β-glucosidase strain is selected from any one of Lactobacillus plantarum LP115, Lactobacillus plantarum subsp. plantarum ATCC14917 and Lactobacillus fermentum CECT5716. In step 3, the catalyst aid comprises the following raw materials by mass percentage: 1% polysorbate 80, 1.5% β-cyclodextrin, 0.4% citric acid, 0.3% sodium citrate, 0.4% calcium chloride, 1% β-glucosidase, 1% β-xylosidase, 4.4% microcrystalline cellulose and 90% purified water; The mass ratio of the pretreated yeast, catalyst, and anhydrous ethanol is 10:1.5:

1.

2. The extraction method of a highly active ginseng extract according to claim 1, characterized in that, In step 2, the parameters of the high-voltage pulsed electric field are set as follows: field strength 20-25 kV / cm, pulse frequency 1000 Hz, time 10-15 min, and temperature 40℃.

3. The extraction method for a highly active ginseng extract according to claim 1, characterized in that, In step 3, the parameters of the supercritical CO2 extraction process are set as follows: extraction pressure 25-35 MPa, CO2 flow rate 150 L / h, extraction temperature 45-55℃, and extraction time 1-2 h.

4. The extraction method for a highly active ginseng extract according to claim 3, characterized in that, In step 4, the ultrafiltration centrifugation process uses an ultrafiltration tube with a molecular weight of 10 kDa; In step 5, the decolorization process uses a decolorizing resin column.

Images