A Ginkgo biloba extract for improving cognitive impairment, its preparation method and application

Abstract

This invention belongs to the field of pharmaceutical technology, specifically relating to a ginkgo leaf extract for improving cognitive impairment, its preparation method, and its application. The method includes the following steps: (1) first, pulverizing and sieving ginkgo leaves to obtain ginkgo leaf powder; (2) then mixing the ginkgo leaf powder with a eutectic solvent and extracting to obtain an extract; (3) finally, passing the extract sequentially through a macroporous resin and a polyamide column for adsorption, collecting the eluent, and obtaining the final product. The eutectic solvent is a mixture of L-carnitine, urea, and trifluoromethanesulfonic acid in a molar ratio of 1.5-2:1:0.2-0.4. This invention uses a specific eutectic solvent in the preparation of the ginkgo leaf extract. Through the synergistic effect of L-carnitine, urea, and trifluoromethanesulfonic acid, the pharmacological activity and the content of the effective components—total flavonol glycosides, total terpene lactones, and proanthocyanidins—are preserved to the greatest extent possible. Furthermore, it requires less equipment and has lower costs.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical technology, specifically relating to a ginkgo leaf extract for improving cognitive impairment, its preparation method, and its application. Background Technology

[0002] Ginkgo biloba, a plant of the Ginkgoaceae family, has leaves and fruits with significant medicinal value. Ginkgo leaf extract is a mixture with unique pharmacological activities extracted from ginkgo leaves. Many scholars both domestically and internationally have used modern separation techniques and testing methods to study the chemical composition of ginkgo leaf extract, identifying over 100 chemical components, including flavonoids, terpene lactones, polyisoprene alcohols, organic acids, alkylphenols and alkylphenolic acids, steroidal compounds, amino acids, and trace elements. Ginkgo leaf extract possesses considerable antioxidant activity, capable of scavenging excess free radicals in the body, preventing lipid peroxidation, enhancing immunity, and delaying aging.

[0003] Chinese invention patent application 201610149596.0 discloses a preparation process for water-soluble ginkgo leaf extract and the water-soluble ginkgo leaf extract itself. The preparation process of the water-soluble ginkgo leaf extract is characterized by using fat-soluble ginkgo leaf extract as raw material and 22-28% hot ethanol as extraction solvent. The process is efficient, safe, simple, and economical. The resulting water-soluble extract is 100% soluble in water. Its effective components include a total flavonoid content of 24-27%, a lactone content of 6-8%, and ginkgolic acid content of less than 5 ppm. The oral liquid prepared with it is pure, transparent, and free of precipitation and suspended matter. It is also easily absorbed by human tissues, and its bioavailability is at least 20% higher than that of ordinary solid preparations.

[0004] Another Chinese invention patent application, 201711095244.2, discloses a method for preparing ginkgo leaf extract. The method uses an ethanol-water solution as the extraction solvent, employing a three-cycle extraction process followed by evaporation to collect the residual alcohol. During the intervals between the three extractions, the ginkgo leaves are subjected to appropriate microwave radiation, which preheats and expands the cells in the effective parts of the leaves. During the subsequent ethanol extraction, these cells rupture prematurely, resulting in a more complete release of the effective substances. The extract is then concentrated in two stages, purified by water precipitation, and further separated, deacidified, concentrated, and dried using a resin column. This yields a ginkgo leaf extract with the removal of harmful total ginkgolic acid and improved purity of total flavonol glycosides and terpene lactones. The resin column is used as the adsorbent, and the ethanol-water solution as the eluent. The advantages of this method include low cost, good separation effect, removal of harmful components, high yield, low pollution, good quality, and ease of industrial production.

[0005] Modern medical research indicates that the pathogenesis of memory and mild cognitive impairment is extremely complex, involving multiple gene defects, environmental influences, neurotoxins, apoptosis, oxidative stress, β-AP (β-amyloid protein) aggregation, inflammatory responses, and calcium deficiency.2+ The disease involves multiple aspects and stages, including overload and degenerative changes in multiple systems such as cholinergic disorders. There are no specific and effective drugs for the symptoms of memory and mild cognitive impairment. Currently, the market mainly uses drugs that improve Alzheimer's disease, such as cholinesterase inhibitors, glutamate receptor antagonists, calcium channel blockers, and estrogens. These drugs are primarily chemical drugs and have significant toxic side effects.

[0006] Therefore, there is an urgent need to provide a ginkgo leaf extract that is high in active ingredients, environmentally friendly, and suitable for industrial production, for improving cognitive impairment. Summary of the Invention

[0007] This invention addresses the problems existing in the prior art by providing a ginkgo leaf extract for improving cognitive impairment, its preparation method, and its application.

[0008] A method for preparing a ginkgo leaf extract that improves cognitive impairment includes the following steps:

[0009] (1) First, crush the ginkgo leaves and sieve them to obtain ginkgo leaf powder;

[0010] (2) Ginkgo leaf powder is then mixed with a eutectic solvent and extracted to obtain an extract;

[0011] (3) Finally, the extract is sequentially passed through a macroporous resin and a polyamide column for adsorption, and the eluent is collected to obtain the final product.

[0012] The eutectic solvent is a mixture of L-carnitine, urea, and trifluoromethanesulfonic acid in a molar ratio of 1.5-2:1:0.2-0.4.

[0013] Preferably, the eutectic solvent is a mixture of L-carnitine, urea, and trifluoromethanesulfonic acid in a molar ratio of 2:1:0.2.

[0014] Preferably, the mesh size of the sieve in step (1) is 60-80 mesh.

[0015] Preferably, the eutectic solvent has a water content of 30-40%.

[0016] Preferably, the mass-to-volume ratio of ginkgo leaf powder to solvent in step (2) is 1 g: 20-30 mL.

[0017] Preferably, the mixing in step (2) requires ultrasonication, the ultrasonic temperature is 30-40℃, the ultrasonic power is 80-100 W, the ultrasonic time is 20-30 min, and centrifugation and filtration are performed after the ultrasonication is completed.

[0018] Preferably, the macroporous resin in step (3) is DM-130, and the adsorption includes adsorbing the extract through DM-130 macroporous resin, first eluting with water until neutral, then eluting with 10-20 column volumes of ethanol solution with a volume fraction of 75-85%, collecting the eluent, concentrating under reduced pressure to obtain a concentrate; then passing the concentrate through a polyamide column, first eluting with 8-10 column volumes of ethanol solution with a volume fraction of 20-30%, and then eluting with 8-10 column volumes of ethanol solution with a volume fraction of 40-50%.

[0019] Preferably, the elution flow rate is 1-5 BV / h and the elution temperature is 15-35℃.

[0020] Preferably, after collecting the eluent in step (3), the eluent needs to be concentrated and dried. The drying is vacuum drying, freeze drying or spray drying, until the water content is 10-15%.

[0021] The present invention also provides a ginkgo leaf extract prepared by the above preparation method, wherein the effective components of the ginkgo leaf extract include total flavonol glycosides, total terpene lactones and proanthocyanidins, and the content ratio of total flavonol glycosides, total terpene lactones and proanthocyanidins is 10-11:3-4:2-3.

[0022] The present invention also provides a pharmaceutical composition for improving cognitive impairment, comprising the above-described ginkgo leaf extract or the ginkgo leaf extract prepared by the above-described preparation method.

[0023] The present invention also provides the use of the above-described Ginkgo biloba extract or the Ginkgo biloba extract prepared by the above-described preparation method or the above-described pharmaceutical composition in the preparation of a medicament for the prevention / treatment and improvement of cognitive impairment.

[0024] Compared with the prior art, the present invention has the following beneficial effects:

[0025] (1) In the preparation of Ginkgo biloba extract, a specific eutectic solvent was used. Through the synergistic effect of L-carnitine, urea and trifluoromethanesulfonic acid, the pharmacological activity and the content of effective components total flavonol glycosides, total terpene lactones and proanthocyanidins were preserved to the greatest extent.

[0026] (2) The present invention uses a specific resin purification process in the preparation of Ginkgo biloba extract, which makes the separation efficiency of effective components high, the separation speed fast, and the experimental cycle short.

[0027] (3) The method for preparing Ginkgo biloba extract provided by the present invention includes ultrasonic dissolution and resin purification, which has low requirements for instruments and equipment, low cost, and is suitable for industrial production.

[0028] (4) The present invention provides a ginkgo leaf extract for improving cognitive impairment, wherein the ratio of total flavonol glycosides, total terpene lactones and proanthocyanidins in the extract is 10-11:3-4:2-3, which can significantly improve cognitive impairment symptoms. Detailed Implementation

[0029] It is worth noting that the raw materials used in this invention are all commercially available products.

[0030] Example 1

[0031] A method for preparing Ginkgo biloba extract that improves cognitive impairment, comprising the following steps:

[0032] (1) Ginkgo leaves are crushed and passed through an 80-mesh sieve to obtain ginkgo leaf powder;

[0033] (2) Then, 5 g of ginkgo leaf powder was ultrasonically mixed with 125 mL of eutectic solvent with a water content of 40% (the eutectic solvent was a mixture of L-carnitine, urea and trifluoromethanesulfonic acid with a molar ratio of 2:1:0.2). The ultrasonic temperature was set to 40℃, the ultrasonic time was 25 min, and the ultrasonic power was 100 W. Then, it was centrifuged at 2000 rpm for 15 min and filtered to obtain ginkgo leaf extract.

[0034] (3) The ginkgo extract was then adsorbed through DM-130 macroporous resin. It was first eluted with water until neutral, and then eluted with 15 column volumes of 80% ethanol solution at a flow rate of 3 BV / h. The eluent was collected and concentrated under reduced pressure to obtain a concentrate. The concentrate was then passed through a polyamide column. It was first eluted with 10 column volumes of 25% ethanol solution at a flow rate of 5 BV / h, and then eluted with 10 column volumes of 45% ethanol solution at a flow rate of 2 BV / h. The concentrate was concentrated and freeze-dried until the water content was 10% to obtain the ginkgo leaf extract.

[0035] Example 2

[0036] A method for preparing Ginkgo biloba extract that improves cognitive impairment, comprising the following steps:

[0037] (1) Ginkgo leaves are crushed and passed through an 80-mesh sieve to obtain ginkgo leaf powder;

[0038] (2) Then, mix 5 g of ginkgo leaf powder with 100 mL of eutectic solvent with a water content of 30% by ultrasonication (the eutectic solvent is a mixture of L-carnitine, urea and trifluoromethanesulfonic acid with a molar ratio of 1.5:1:0.4). Set the ultrasonic temperature to 30℃, the ultrasonic time to 20 min, and the ultrasonic power to 80 W. Then, centrifuge at 2000 rpm for 15 min, filter, and obtain ginkgo leaf extract.

[0039] (3) The ginkgo extract was then adsorbed through DM-130 macroporous resin. It was first eluted with water until neutral, and then eluted with 10 column volumes of 75% ethanol solution at a flow rate of 4 BV / h. The eluent was collected and concentrated under reduced pressure to obtain a concentrate. The concentrate was then passed through a polyamide column. It was first eluted with 8 column volumes of 25% ethanol solution at a flow rate of 5 BV / h, and then eluted with 8 column volumes of 45% ethanol solution at a flow rate of 1 BV / h. The concentrate was concentrated and freeze-dried until the water content was 12% to obtain the ginkgo leaf extract.

[0040] Example 3

[0041] A method for preparing Ginkgo biloba extract that improves cognitive impairment, comprising the following steps:

[0042] (1) Ginkgo leaves are crushed and passed through a 60-mesh sieve to obtain ginkgo leaf powder;

[0043] (2) Then, 5 g of ginkgo leaf powder was ultrasonically mixed with 150 mL of eutectic solvent with a water content of 40% (the eutectic solvent was a mixture of L-carnitine, urea and trifluoromethanesulfonic acid with a molar ratio of 2:1:0.2). The ultrasonic temperature was set to 40℃, the ultrasonic time was 30 min, and the ultrasonic power was 100 W. Then, it was centrifuged at 2000 rpm for 20 min and filtered to obtain ginkgo leaf extract.

[0044] (3) The ginkgo extract was then adsorbed through DM-130 macroporous resin. It was first eluted with water until neutral, and then eluted with 20 column volumes of 85% ethanol solution at a flow rate of 3 BV / h. The eluent was collected and concentrated under reduced pressure to obtain a concentrate. The concentrate was then passed through a polyamide column. It was first eluted with 10 column volumes of 25% ethanol solution at a flow rate of 5 BV / h, and then eluted with 10 column volumes of 45% ethanol solution at a flow rate of 2 BV / h. The concentrate was concentrated and freeze-dried until the water content was 15% to obtain the ginkgo leaf extract.

[0045] Comparative Example 1

[0046] A method for preparing a ginkgo leaf extract that improves cognitive impairment differs from Example 1 only in that the eutectic solvent in step (2) is a mixture of urea and trifluoromethanesulfonic acid in a molar ratio of 1:0.2.

[0047] A method for preparing Ginkgo biloba extract that improves cognitive impairment, comprising the following steps:

[0048] (1) Ginkgo leaves are crushed and passed through an 80-mesh sieve to obtain ginkgo leaf powder;

[0049] (2) Then, 5 g of ginkgo leaf powder was ultrasonically mixed with 125 mL of eutectic solvent with a water content of 40% (the eutectic solvent is a mixture of urea and trifluoromethanesulfonic acid with a molar ratio of 1:0.2). The ultrasonic temperature was set to 40℃, the ultrasonic time was 25 min, and the ultrasonic power was 100 W. Then, it was centrifuged at 2000 rpm for 15 min and filtered to obtain ginkgo leaf extract.

[0050] (3) The ginkgo extract was then adsorbed through DM-130 macroporous resin. It was first eluted with water until neutral, and then eluted with 15 column volumes of 80% ethanol solution at a flow rate of 3 BV / h. The eluent was collected and concentrated under reduced pressure to obtain a concentrate. The concentrate was then passed through a polyamide column. It was first eluted with 10 column volumes of 25% ethanol solution at a flow rate of 5 BV / h, and then eluted with 10 column volumes of 45% ethanol solution at a flow rate of 2 BV / h. The concentrate was concentrated and freeze-dried until the water content was 10% to obtain the ginkgo leaf extract.

[0051] Comparative Example 2

[0052] A method for preparing a ginkgo leaf extract that improves cognitive impairment differs from Example 1 only in that the eutectic solvent in step (2) is a mixture of L-carnitine and trifluoromethanesulfonic acid in a molar ratio of 2:0.2.

[0053] A method for preparing Ginkgo biloba extract that improves cognitive impairment, comprising the following steps:

[0054] (1) Ginkgo leaves are crushed and passed through an 80-mesh sieve to obtain ginkgo leaf powder;

[0055] (2) Then, mix 5 g of ginkgo leaf powder with 125 mL of eutectic solvent with a water content of 40% by ultrasonication (the eutectic solvent is a mixture of L-carnitine and trifluoromethanesulfonic acid with a molar ratio of 2:0.2). Set the ultrasonic temperature to 40℃, the ultrasonic time to 25 min, and the ultrasonic power to 100 W. Then, centrifuge at 2000 rpm for 15 min, filter, and obtain ginkgo leaf extract.

[0056] (3) The ginkgo extract was then adsorbed through DM-130 macroporous resin. It was first eluted with water until neutral, and then eluted with 15 column volumes of 80% ethanol solution at a flow rate of 3 BV / h. The eluent was collected and concentrated under reduced pressure to obtain a concentrate. The concentrate was then passed through a polyamide column. It was first eluted with 10 column volumes of 25% ethanol solution at a flow rate of 5 BV / h, and then eluted with 10 column volumes of 45% ethanol solution at a flow rate of 2 BV / h. The concentrate was concentrated and freeze-dried until the water content was 10% to obtain the ginkgo leaf extract.

[0057] Comparative Example 3

[0058] A method for preparing a ginkgo leaf extract that improves cognitive impairment differs from Example 1 only in that the eutectic solvent in step (2) is a mixture of L-carnitine and urea in a molar ratio of 2:1.

[0059] A method for preparing Ginkgo biloba extract that improves cognitive impairment, comprising the following steps:

[0060] (1) Ginkgo leaves are crushed and passed through an 80-mesh sieve to obtain ginkgo leaf powder;

[0061] (2) Then, mix 5 g of ginkgo leaf powder with 125 mL of eutectic solvent with a water content of 40% by ultrasonication (the eutectic solvent is a mixture of L-carnitine and urea with a molar ratio of 2:1). Set the ultrasonic temperature to 40℃, the ultrasonic time to 25 min, and the ultrasonic power to 100 W. Then, centrifuge at 2000 rpm for 15 min, filter, and obtain ginkgo leaf extract.

[0062] (3) The ginkgo extract was then adsorbed through DM-130 macroporous resin. It was first eluted with water until neutral, and then eluted with 15 column volumes of 80% ethanol solution at a flow rate of 3 BV / h. The eluent was collected and concentrated under reduced pressure to obtain a concentrate. The concentrate was then passed through a polyamide column. It was first eluted with 10 column volumes of 25% ethanol solution at a flow rate of 5 BV / h, and then eluted with 10 column volumes of 45% ethanol solution at a flow rate of 2 BV / h. The concentrate was concentrated and freeze-dried until the water content was 10% to obtain the ginkgo leaf extract.

[0063] Comparative Example 4

[0064] A method for preparing a ginkgo leaf extract that improves cognitive impairment differs from Example 1 only in that ultrasound is not performed in step (2).

[0065] A method for preparing Ginkgo biloba extract that improves cognitive impairment, comprising the following steps:

[0066] (1) Ginkgo leaves are crushed and passed through an 80-mesh sieve to obtain ginkgo leaf powder;

[0067] (2) Mix 5 g of ginkgo leaf powder with 125 mL of eutectic solvent with a water content of 40% (the eutectic solvent is a mixture of L-carnitine, urea and trifluoromethanesulfonic acid with a molar ratio of 2:1:0.2, then centrifuge at 2000 rpm for 15 min, filter, and obtain ginkgo leaf extract.

[0068] (3) The ginkgo extract was then adsorbed through DM-130 macroporous resin. It was first eluted with water until neutral, and then eluted with 15 column volumes of 80% ethanol solution at a flow rate of 3 BV / h. The eluent was collected and concentrated under reduced pressure to obtain a concentrate. The concentrate was then passed through a polyamide column. It was first eluted with 10 column volumes of 25% ethanol solution at a flow rate of 5 BV / h, and then eluted with 10 column volumes of 45% ethanol solution at a flow rate of 2 BV / h. The concentrate was concentrated and freeze-dried until the water content was 10% to obtain the ginkgo leaf extract.

[0069] Comparative Example 5

[0070] A method for preparing a ginkgo leaf extract that improves cognitive impairment differs from Example 1 only in that, in step (3), DM-130 macroporous resin is replaced with HP-20 macroporous resin.

[0071] A method for preparing Ginkgo biloba extract that improves cognitive impairment, comprising the following steps:

[0072] (1) Ginkgo leaves are crushed and passed through an 80-mesh sieve to obtain ginkgo leaf powder;

[0073] (2) Then, 5 g of ginkgo leaf powder was ultrasonically mixed with 125 mL of eutectic solvent with a water content of 40% (the eutectic solvent was a mixture of L-carnitine, urea and trifluoromethanesulfonic acid with a molar ratio of 2:1:0.2). The ultrasonic temperature was set to 40℃, the ultrasonic time was 25 min, and the ultrasonic power was 100 W. Then, it was centrifuged at 2000 rpm for 15 min and filtered to obtain ginkgo leaf extract.

[0074] (3) The ginkgo extract was then adsorbed onto an HP-20 macroporous resin. It was first eluted with water until neutral, and then eluted with 15 column volumes of 80% ethanol solution at a flow rate of 3 BV / h. The eluent was collected and concentrated under reduced pressure to obtain a concentrate. The concentrate was then passed through a polyamide column. It was first eluted with 10 column volumes of 25% ethanol solution at a flow rate of 5 BV / h, and then eluted with 10 column volumes of 45% ethanol solution at a flow rate of 2 BV / h. The concentrate was concentrated and freeze-dried until the water content was 10% to obtain the ginkgo leaf extract.

[0075] Content determination

[0076] The contents of total flavonol glycosides, total terpene lactones, and proanthocyanidins in the Ginkgo biloba extracts prepared in Examples 1-3 and Comparative Examples 1-5 were determined according to the content determination method of Ginkgo biloba extract in the 2020 edition of the Chinese Pharmacopoeia. The contents of the three were obtained and the results are shown in Table 1 below.

[0077] Table 1. Determination of total flavonol glycosides, total terpene lactones and proanthocyanidins in Ginkgo biloba leaf extract.

[0078]

[0079] Experiment 1: The ameliorative effect of the Ginkgo biloba extract of this invention on Alzheimer's disease model mice.

[0080] 1. Test samples: Ginkgo biloba extracts prepared in Examples 1-3 and Comparative Examples 1-5 of this invention.

[0081] 2. Experimental animals: APP / PS1 transgenic mice, SPF grade males, 6 months old, weighing 20±2 g, purchased from Guangdong Provincial Medical Laboratory Animal Center, with free access to water and food, room temperature maintained at 24±2℃, and relative humidity 55-65%.

[0082] 3. Grouping: Mice were randomly divided into 10 groups, namely the control group, Example 1 (low, medium and high dose groups), Example 2-3 and Comparative Examples 1-5, with 10 mice in each group. Example 1-3 and Comparative Examples 1-5 were administered the corresponding products by gavage, while the control group was administered an equal volume of physiological saline by gavage. The dosages are shown in Table 2. Mice were administered the products twice by gavage, with an interval of 2 hours, for 7 consecutive days.

[0083] 4. Experimental Methods: After administering the drug, mice in each group were placed in a box to acclimatize for 30 minutes. Then, a 36V AC current was applied. Upon receiving the electric shock, the mice jumped onto a rubber platform to avoid the noxious stimulus. If a mouse jumped off the platform, it would return to the platform after being stimulated again. This training was repeated for 5 minutes until the mice developed memory and remained on the platform. Testing was conducted 24 hours after training. The mice were placed on the platform, and the latency to jump off the platform for the first time and the number of times they jumped off the platform within 5 minutes (i.e., the number of errors) were recorded.

[0084] 5. Experimental Results: As shown in Table 2, compared with the control group, the number of errors in the low-dose group of Ginkgo biloba extract in Example 1 was reduced ( P <0.05%, the number of errors was significantly reduced in the high and medium dose groups of Ginkgo biloba extract (Ginkgo biloba extract <0.05). P <0.01), the latency period was significantly prolonged in the high, medium and low dose groups ( P <0.01).

[0085] Compared with Comparative Examples 1-5, at the same dosage, the number of errors in Examples 1-3 was significantly lower than that in the comparative examples, and the latency period was significantly longer than that in the comparative examples. This indicates that the Ginkgo biloba extract prepared in this invention can consolidate the memory of mice in the passive avoidance experiment and enhance the learning ability of mice.

[0086] Table 2. Effects of Ginkgo biloba extract on learning and memory in mice ( ±s, n=10)

[0087]

[0088] Note: * This indicates a comparison with the control group. P <0.05, ** This indicates a comparison with the control group. P <0.01.

[0089] Finally, it should be noted that the above content is only used to illustrate the technical solution of the present invention, and is not intended to limit the scope of protection of the present invention. Simple modifications or equivalent substitutions made by those skilled in the art to the technical solution of the present invention do not depart from the essence and scope of the technical solution of the present invention.

Claims

1. A method for preparing ginkgo biloba leaf extract for improving cognitive impairment, characterized by, The method comprises the following steps: (1) crushing ginkgo leaves, sieving, and obtaining ginkgo leaf powder; (2) mixing the ginkgo leaf powder with a eutectic solvent, extracting, and obtaining an extract; (3) finally, sequentially adsorbing the extract through a macroporous resin column and a polyamide column, collecting the eluate, and obtaining the product, wherein the eutectic solvent is a mixture of L-carnitine, urea and trifluoromethanesulfonic acid with a molar ratio of 1.5-2:1:0.2-0.

4.

2. The production method according to claim 1, characterized by, The mesh number of the sieving in step (1) is 60-80.

3. The preparation method according to claim 1, characterized in that, The water content of the eutectic solvent is 30-40%.

4. The production method according to claim 3, characterized by, The mass-volume ratio of the ginkgo leaf powder to the solvent in step (2) is 1 g:20-30 mL.

5. The preparation method according to claim 1, characterized in that, The mixing in step (2) needs ultrasonic treatment, the temperature of the ultrasonic treatment is 30-40℃, the power of the ultrasonic treatment is 80-100 W, the time of the ultrasonic treatment is 20-30 min, and centrifugation and filtration are performed after the ultrasonic treatment.

6. The production method according to claim 5, wherein The macroporous resin in step (3) is DM-130, and the adsorption comprises adsorbing the extract through a DM-130 macroporous resin column, eluting with water to neutral, eluting with 10-20 times the column volume of an ethanol solution with a volume fraction of 75-85%, collecting the eluate, concentrating under reduced pressure, obtaining a concentrated solution, and then passing the concentrated solution through a polyamide column, eluting with 8-10 times the column volume of an ethanol solution with a volume fraction of 20-30%, eluting with 8-10 times the column volume of an ethanol solution with a volume fraction of 40-50%, and the flow rate of the elution is 1-5 BV / h, and the temperature of the elution is 15-35℃.

7. The preparation method according to claim 1, characterized in that, After the collection of the eluate in step (3) is completed, the eluate needs to be concentrated and dried, the drying is vacuum drying, freeze-drying or spray drying, and the water content after the drying is 10-15%.

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