A method for extracting and purifying huperzine A

Abstract

The present invention relates to a method for extracting and purifying huperzine A. The method comprises: (1) cutting melaleuca alternifolia into sections and drying; (2) adding alkaline ethanol, stirring and impregnating, then heating and refluxing for extraction under stirring conditions, filtering, and concentrating; (3) stirring, then cooling and refrigerating, and filtering; (4) concentrating to an extract, adding an acidic aqueous solution, filtering, adding an alkaline aqueous solution, and rotating to dryness; (6) adding a mixed solvent of ethyl acetate and n-heptane, stirring, adding an alkaline aqueous solution for extraction, discarding the aqueous solution, crystallizing, filtering, and drying to obtain huperzine A. The method has the advantages of low cost, low organic solvent usage, short production cycle, and easy operation, and is suitable for industrial production. The obtained huperzine A has a high yield and purity.

Description

Technical Field

[0001] The invention belongs to the field of traditional Chinese medicine extraction methods, and particularly relates to a method for extracting and purifying huperzine A. Background Art

[0002] The whole plant of the plant, also known as snake's foot grass, snake's twig, Qianjinzha, dwarf fir, Wannian fir, iron plate grass, Qianjin worm, and Jinbuhuan, is harvested in late summer and early autumn, de-soiled, and sun-dried. The spores are collected between July and August and dried for medicinal use. It is widely distributed in Northeast my country, the Yangtze River Basin, Zhejiang, Fujian, Guangdong, Sichuan, Guizhou, and Yunnan. Qiancengta is known for its heat-clearing and detoxifying properties, tissue-stimulating and hemostatic properties, and stasis-dispersing and swelling-reducing properties. It is used to treat injuries from falls, congestion, swelling, and internal bleeding. It can also be used externally to treat carbuncles, snake bites, burns, and scalds. Recent clinical studies have shown that extracts from this plant are effective in treating schizophrenia, myasthenia gravis, and senile dementia, as well as improving memory.

[0003] Huperzine A, a highly important natural plant alkaloid found in the genus Melaleuca, is a potent, reversible, and highly selective second-generation acetylcholinesterase inhibitor. Its high lipid solubility and small molecular weight allow it to penetrate the blood-brain barrier effectively. Once inside the central nervous system, it is distributed primarily in the frontal and temporal lobes, hippocampus, and other brain regions closely associated with learning and memory. At low doses, it exhibits a potent inhibitory effect on acetylcholinesterase (AchE), significantly increasing acetylcholine (Ach) levels in the synaptic cleft within these distribution areas. This enhances neuronal excitation and strengthens the excitation of learning and memory brain regions, improving cognitive function, enhancing memory retention, and facilitating memory retrieval. It is currently the most successfully developed drug in China for the treatment of Alzheimer's disease (senile dementia). Huperzine A is also used for myasthenia gravis, senile memory loss, Alzheimer's disease, learning disabilities, and memory enhancement in adolescents. It also possesses anti-cancer, anti-inflammatory, and HIV-inhibiting properties.

[0004] Currently, there is considerable research on the extraction and separation of Huperzine A from Psoralea cochinchinensis. Chinese patent CN104262251B discloses a method for extracting Huperzine A from Psoralea cochinchinensis. The method comprises pulverizing the Psoralea cochinchinensis, performing ultrasonic countercurrent extraction, membrane separation and purification, concentration, acid dissolution, extraction, concentration, crystallization and recrystallization, and drying to obtain the Huperzine A product. However, due to the use of ultrasonic countercurrent extraction equipment, extraction, multiple crystallizations and recrystallizations, and membrane filtration during the process, the method involves multiple production steps, complex operations, significant process losses, high organic solvent consumption, low yield, and high cost. Chinese patent CN102617469A discloses a method for extracting Huperzine A from Melaleuca alternifolia, which comprises crushing Melaleuca alternifolia medicinal materials, ultrasonic extraction with acid solution, concentration, pH adjustment with ammonia water, extraction with chloroform, redissolving with acid solution, filtering with activated carbon for impurity removal, adjusting pH with ammonia water for a second time, extraction with chloroform again, crystallization and recrystallization, and finally drying to obtain a product, wherein the purity of Huperzine A in the product reaches 99%, and the yield of Huperzine A is about 0.016%. Due to the multiple extractions in the process, the loss is large, the yield is low, and the cost is high. Chinese patent CN101693689A discloses a new method for extracting and separating high-purity Huperzine A from Melaleuca alternifolia, which comprises crushing Melaleuca alternifolia medicinal materials appropriately, extracting with acid solution, adjusting pH with ammonia water, separation by adsorption with macroporous resin, purification by high-speed countercurrent chromatography, and concentration and drying to obtain a finished product, wherein the purity of Huperzine A in the finished product can reach more than 98%, but the high-speed countercurrent chromatography equipment investment is large and the processing capacity is limited during the process, making it difficult to adapt to industrial production. Chinese patent CN102491946A discloses a method for separating and purifying Huperzine A using molecular imprinting technology. This method uses ultrasonic countercurrent extraction on the medicinal material of Melaleuca alternifolia, and after concentration and decolorization, molecular imprinting polymerization, crystallization and recrystallization can produce a product with a purity of more than 99%. This inventive method directly uses ultrasonic countercurrent extraction on the medicinal material. Since the active ingredient content in Melaleuca alternifolia is extremely low, the extracted impurities are relatively large, which to a certain extent hinders the transfer of the active substance in the solvent, affects the extraction yield of Huperzine A, and is not conducive to subsequent purification processes. In addition, the molecular imprinting technology used in this method has high operational requirements, complex control, and immature technology. It is only suitable for small-batch production and is not conducive to scale-up and industrial production. Therefore, it is necessary to explore a new method for extracting Huperzine A so that Huperzine A has both good yield and purity and is suitable for scale-up and industrial production. Summary of the Invention

[0005] The object of the present invention is to provide an extraction and purification method suitable for industrial production, which can obtain high-purity huperzine A with a yield of more than 75%.

[0006] The present invention provides a method for extracting and purifying huperzine A, comprising the following steps:

[0007] (1) Cut the Thousand Layer Pagoda into 2-3 cm segments and dry them;

[0008] (2) adding alkaline ethanol to the dried melaleuca in step (1), stirring and impregnating, then heating and refluxing under stirring, filtering, and concentrating to an ethanol concentration of 60%-65%;

[0009] (3) stirring the concentrated solution in step (2), cooling it to below 10° C., refrigerating it, and filtering it;

[0010] (4) concentrating the filtered solution in step (3) to an extract, adding an acidic aqueous solution to adjust the pH value to 2.8-3.0, filtering, adjusting the pH value of the filtered solution to 8.5-9.5 with an alkaline aqueous solution, and then drying the water to obtain a solid;

[0011] (5) adding a mixed solvent of ethyl acetate and n-heptane to the solid matter in step (4), stirring at 60-70° C., then adding an alkaline aqueous solution with a pH value of 8.5-9.5 at 60-70° C. and stirring, keeping the temperature and standing to separate, discarding the aqueous solution, standing and cooling the obtained organic solution to 15-25° C. for crystallization, filtering, and drying to obtain huperzine A, wherein the volume ratio of ethyl acetate to n-heptane in the mixed solvent of ethyl acetate and n-heptane is 1:(2-3), and the mass ratio of the mixed solvent of ethyl acetate and n-heptane to the solid matter is (3-5):1.

[0012] Preferably, in step (1), the Thousand Layer Pagoda is first sorted and cleaned before being cut into 2-3 cm segments. The purpose is to sort the Thousand Layer Pagoda medicinal materials to remove non-medicinal impurities before alcohol is added, and then wash with water to remove impurities such as mud, dust, etc., which reduces the difficulty of subsequent processing and is conducive to the purification of the target component.

[0013] Preferably, in step (2), the ratio of alkaline ethanol to dried melaleuca alternifolia is (7-10) mL:1 g; the concentration of alkaline ethanol is 90-95%, and the pH value is 8.5-9.5.

[0014] Preferably, in step (2), the stirring and immersion temperature is 40-45° C., and the stirring and immersion time is 2-4 hours; the heating and reflux extraction times are 3-5 times, and the extraction time each time is 2-4 hours.

[0015] Preferably, in step (3), the stirring temperature is room temperature, the stirring time is 8 to 15 minutes, and the standing refrigeration time is more than 24 hours.

[0016] Preferably, the acidic aqueous solution in step (4) is a hydrochloric acid solution or a sulfuric acid solution; and the alkaline aqueous solution is a sodium hydroxide solution, a potassium hydroxide solution or ammonia water.

[0017] Preferably, the stirring time in step (5) is 10-20 min at 60-70°C.

[0018] Preferably, in step (5), the stirring time of adding the alkaline aqueous solution with a pH value of 8.5 to 9.5 at 60-70° C. is 20-40 minutes.

[0019] Preferably, in step (5), the ratio of the alkaline aqueous solution with a pH value of 8.5 to 9.5 to the solid matter is (8-12) mL:1 g.

[0020] Preferably, the extraction times in step (5) are 3 to 4 times. The extraction here refers to adding an alkaline aqueous solution with a pH value of 8.5 to 9.5 under the above-mentioned conditions of 60-70°C, stirring, keeping the temperature and standing to separate the layers, and discarding the aqueous solution.

[0021] The alkaline ethanol involved in the present invention is prepared by adding an alkaline substance, such as sodium hydroxide solution, ammonia water, etc., into an ethanol solution.

[0022] Beneficial effects

[0023] The present invention has the advantages of low cost, low organic solvent consumption, short production cycle, and easy operation, and is suitable for industrial production. The production cost is about 420-460 yuan / g (production cost=total production cost divided by the weight of the product huperzine A), and the organic solvent consumption is about 7.0-8.7 L / g (organic solvent consumption=total volume of organic solvent consumption divided by the weight of the product huperzine A).

[0024] The product Huperzine A obtained by the present invention has high purity (HPLC test purity is greater than 99%) and a yield of greater than 75% (Huperzine A yield = final Huperzine A product mass / actual Huperzine A mass contained in the melaleuca tower × 100%). DETAILED DESCRIPTION

[0025] Below in conjunction with specific embodiment, further set forth the present invention.Should be understood that these embodiments are only used to illustrate the present invention and are not used in limiting the scope of the present invention.In addition, should be understood that after reading the content taught by the present invention, those skilled in the art can make various changes or modifications to the present invention, and these equivalent forms fall equally within the scope limited by the appended claims of the application.

[0026] Example 1

[0027] A method for extracting and purifying huperzine A comprises:

[0028] (1) Pretreatment: Take the dried Chinese angelica root (the mass content of huperzine A is 0.05%), remove impurities, wash it with water, cut it into 2-3 cm segments, dry it, and take 20 kg of dried Chinese angelica root for subsequent steps.

[0029] (2) Alcohol extraction and concentration: The pre-treated melaleuca extract was placed in an extraction tank, and 160 L of alkaline ethanol with a volume fraction of 90% and a pH value of 8.95 was added. The extract was stirred and immersed at 40°C for 4 hours, and then heated under reflux for extraction. The extraction was performed 4 times, each time for 2 hours, and filtered. The filtrate was concentrated to an alcohol concentration of 60% each time, and the concentrated solution was collected. During the concentration, the vacuum degree was controlled at -0.04 to -0.08 MPa, and the temperature was controlled at 50 to 60°C.

[0030] (3) Alcohol precipitation: The concentrated solution collected in the above four times was transferred to an alcohol precipitation tank and stirred at room temperature for 10 minutes. Low-temperature water was opened and refrigerated at 5-8°C for 30 hours, and then filtered.

[0031] (4) Acid and base adjustment: The filtered solution in step (3) is pumped into a concentrator, and the vacuum is controlled at -0.08 to -0.04 MPa and the temperature is controlled at 50 to 60° C., and the solution is concentrated under reduced pressure to an extract-like state. After cooling to room temperature, the pH value is adjusted to 2.8 with a 1% by volume HCl solution, filtered, and then the pH value is adjusted to 9 with a 40% by volume sodium hydroxide solution. The water is then dried to obtain a solid.

[0032] (5) Extraction and crystallization: The solid obtained in step (4) was added with a mixed solvent of ethyl acetate and n-heptane (the volume ratio of ethyl acetate and n-heptane was 1:2), the mass ratio of the mixed solvent of ethyl acetate and n-heptane to the solid was 4:1, and the mixture was heated to 60-70°C and stirred for 10 min. An aqueous sodium hydroxide solution with a pH value of 9.0 was added, the ratio of the aqueous sodium hydroxide solution to the solid was 10 mL:1 g, and the mixture was stirred at this temperature for 20 min. The mixture was allowed to stand for stratification, and the lower aqueous solution was discarded. The extraction was performed three times according to this method. The obtained organic solution was allowed to stand for cooling and crystallization. The mixture was cooled to 20°C for crystallization for 1.5 hours, filtered, and dried to obtain 7.8 g of white huperzine A with a purity of 99.4% and a yield of 78%.

[0033] The production cost of this embodiment is about 456 yuan / g, and the organic solvent consumption is about 8.6 L / g.

[0034] Example 2

[0035] A method for extracting and purifying huperzine A comprises:

[0036] (1) Pretreatment: Take the shade-dried Herba Coptidis (the mass content of Huperzine A is 0.05%), remove impurities, rinse with water, cut into 2-3 cm segments, dry, and take 30 kg of the dried Herba Coptidis for subsequent steps.

[0037] (2) Alcohol extraction and concentration: put the pre-treated Thousand Layer Pagoda into an extraction tank, add 300L of alkaline ethanol with a volume fraction of 90% and a pH value of 9.0, stir and soak at 45°C for 2 hours, then heat and reflux for extraction, extract 3 times in total, each time for 2 hours, filter, and concentrate the filtrate each time until the alcohol concentration reaches 65%, collect the concentrate, and control the vacuum degree at 45°C during concentration.

[0038] -0.04~-0.08MPa, temperature controlled at 50~60℃.

[0039] (3) Alcohol precipitation: The concentrated solution collected in the above three times was transferred to an alcohol precipitation tank and stirred at room temperature for 15 minutes. Low-temperature water was opened and refrigerated at 4-6°C for 28 hours, and then filtered.

[0040] (4) Acid and base adjustment: The filtered solution in step (3) is pumped into a concentrator, and the vacuum is controlled at -0.08 to -0.04 MPa and the temperature is controlled at 50 to 60° C., and the solution is concentrated under reduced pressure to an extract-like state. After cooling to room temperature, the pH value is adjusted to 3.0 with a 1% by volume HCl solution, filtered, and then the pH value is adjusted to 8.9 with a 40% by volume sodium hydroxide solution. The water is then dried to obtain a solid.

[0041] (5) Extraction and crystallization: The solid obtained in step (4) was added with a mixed solvent of ethyl acetate and n-heptane (the volume ratio of ethyl acetate and n-heptane was 1:3), the mass ratio of the mixed solvent of ethyl acetate and n-heptane to the solid was 3:1, and the mixture was heated to 60-70°C and stirred for 15 minutes. An aqueous sodium hydroxide solution with a pH value of 8.9 was added, and the ratio of the aqueous sodium hydroxide solution to the solid was 8 mL:1 g. The mixture was stirred at this temperature for 30 minutes, and the mixture was allowed to stand for stratification. The lower aqueous solution was discarded, and the extraction was performed four times according to this method. The obtained organic solution was allowed to stand for cooling and crystallization. The mixture was cooled to 15°C for crystallization for 1 hour, filtered, and dried to obtain 12 g of white huperzine A with a purity of 99.2% and a yield of 80%.

[0042] The production cost of this embodiment is about 441 yuan / g, and the organic solvent consumption is about 7.8 L / g.

[0043] Example 3

[0044] A method for extracting and purifying huperzine A comprises:

[0045] (1) Pretreatment: Take the dried radix serrata medicinal material (the mass content of huperzine A is 0.05%), remove impurities, quickly wash it with water, cut it into 2-3 cm segments, dry it, and take 100 kg of the dried radix serrata for subsequent steps.

[0046] (2) Alcohol extraction and concentration: The pre-treated Thousand Layer Tower was placed in an extraction tank, and 700 L of alkaline ethanol with a volume fraction of 90% and a pH value of 9.2 was added. The mixture was stirred and immersed at 45°C for 4 hours, and then heated under reflux for extraction. The extraction was performed 5 times, each time for 4 hours, and filtered. The filtrate was concentrated to an alcohol concentration of 65% each time, and the concentrated solution was collected. During the concentration, the vacuum degree was controlled at 0.04-0.08 MPa, and the temperature was controlled at 50-60°C.

[0047] (3) Alcohol precipitation: The concentrated solution collected from the above five times was transferred to an alcohol precipitation tank and stirred at room temperature for 8 minutes. Low-temperature water was opened and refrigerated at 0-3°C for 25 hours, and then filtered.

[0048] (4) Acid and base adjustment: The filtered solution in step (3) is pumped into a concentrator, and the vacuum is controlled at -0.08 to -0.04 MPa and the temperature is controlled at 50 to 60° C., and the solution is concentrated under reduced pressure to an extract form. After cooling to room temperature, the pH value is adjusted to 2.8 with a 1% by volume HCl solution, filtered, and then the pH value is adjusted to 9.3 with a 40% by volume potassium hydroxide solution. The water is then dried to obtain a solid.

[0049] (5) Extraction and crystallization: The solid obtained in step (4) was added with a mixed solvent of ethyl acetate and n-heptane (the volume ratio of ethyl acetate and n-heptane was 1:3), the mass ratio of the mixed solvent of ethyl acetate and n-heptane to the solid was 5:1, and the mixture was heated to 60-70°C and stirred for 20 minutes. A potassium hydroxide aqueous solution with a pH value of 9.0 was added, and the ratio of the potassium hydroxide aqueous solution to the solid was 12 mL:1 g. The mixture was stirred at this temperature for 40 minutes, and the mixture was allowed to stand for stratification. The lower aqueous solution was discarded, and the extraction was performed four times according to this method. The obtained organic solution was allowed to stand for cooling and crystallization. The mixture was cooled to 25°C for crystallization for 2 hours, filtered, and dried to obtain 42 g of white huperzine A with a purity of 99.5% and a yield of 84%.

[0050] The production cost of this embodiment is about 428 yuan / g, and the organic solvent consumption is about 8.7 L / g.

[0051] Comparative Example 1

[0052] According to Example 1, steps (1) and (2) in Example 1 were repeated, and the concentrated solutions collected four times were combined. The combined solutions were then divided into five groups, each with the same solution volume, and the following experiments were performed respectively:

[0053] Group 1: The concentrated solution was directly subjected to steps (4)-(5) of Example 1 to obtain 1.296 g of huperzine A.

[0054] Group 2: The concentrate was stirred at room temperature for 10 minutes, cooled and refrigerated at 13-16°C for 30 hours with low-temperature water, filtered, and then steps (4)-(5) of Example 1 were repeated to obtain 1.404 g of huperzine A.

[0055] Group 3: Ethanol was added to the concentrate until the alcohol concentration was 70%, and steps (3) to (5) in Example 1 were repeated to obtain 1.428 g of huperzine A.

[0056] Group 4: The concentrate was further concentrated to an alcohol concentration of 55%, and steps (3) to (5) in Example 1 were repeated to obtain 1.272 g of huperzine A.

[0057] Group 5: Repeat steps (3) to (5) of Example 1 with the concentrated solution to obtain 1.548 g of huperzine A.

[0058] The purity, yield, production cost, and organic solvent consumption of Huperzine A in the five experiments are shown in Table 1.

[0059] Table 1

[0060] Huperzine A purity Yield production costs Organic solvent consumption Group 1 87.6％ 65％ About 549 yuan / g About 10.3L / g Group 2 92％ 70.2％ About 507 yuan / g About 9.5L / g Group 3 93.6％ 71.4％ About 498 yuan / g About 9.4L / g Group 4 96.1％ 63.6％ About 559 yuan / g About 10.5L / g Group 5 99.2％ 77.4％ About 460 yuan / g About 8.6L / g

[0061] From this comparative example, it can be seen that the purity and yield of Huperzine A (the fifth group) of the present invention are higher than those of the first, second, third and fourth groups of the control, and it has the advantages of low production cost and low organic solvent consumption.

[0062] Comparative Example 2

[0063] According to Example 2, steps (1) to (4) in Example 2 were repeated, and the solution before drying in step (4) was divided into 9 groups, each with the same volume. 8 groups of solutions were dried to obtain solids, and 1 group of solutions was not dried. The following experiments were performed respectively:

[0064] Group 1: According to Example 2, n-heptane in step (5) was replaced with petroleum ether, and the rest was the same as Example 2 to obtain 1.21 g of huperzine A.

[0065] Group 2: According to Example 2, the volume ratio of ethyl acetate to n-heptane in step (5) was changed to 1:1, and the rest was the same as Example 2, to obtain 1.2 g of huperzine A.

[0066] Group 3: According to Example 2, the volume ratio of ethyl acetate to n-heptane in step (5) was changed to 1:5, and the rest was the same as Example 2, to obtain 1.169 g of huperzine A.

[0067] Group 4: According to Example 2, the mass ratio of the mixed solvent of ethyl acetate and n-heptane to the solid matter in step (5) was changed to 1.5:1, and the rest was the same as Example 2, to obtain 1.0855 g of huperzine A.

[0068] Group 5: According to Example 2, the mass ratio of the mixed solvent of ethyl acetate and n-heptane to the solid matter in step (5) was changed to 7:1, and the rest was the same as Example 2, to obtain 1.19 g of huperzine A.

[0069] Group 6: According to Example 2, the stirring temperature in step (5) was changed to 50° C., and the rest was the same as Example 2, to obtain 1.14 g of huperzine A.

[0070] Group 7: According to Example 2, the stirring temperature in step (5) was changed to 80°C, and the rest was the same as Example 2, to obtain 1.27 g of huperzine A.

[0071] In the eighth group, the alkaline solution was extracted with chloroform three times for 8 minutes each time. The volume of chloroform each time was 4.2 times the volume of the alkaline solution. The chloroform phases were collected and combined, and concentrated under vacuum until dryness. 68 mL of acetone was added to dissolve the obtained dry powder, which was cooled to 8°C and crystallized twice. The extracted powder was dried to obtain 1.002 g of huperzine A.

[0072] Group 9: According to step (5) in Example 2, 1.35 g of huperzine A was obtained.

[0073] The purity, yield, production cost, and organic solvent consumption of Huperzine A in the nine experiments are shown in Table 2.

[0074] Table 2

[0075] Huperzine A purity Yield production costs Organic solvent consumption Group 1 92.6％ 72.5％ About 486 yuan / g About 8.5L / g Group 2 90.9％ 71.9％ About 490 yuan / g About 8.6L / g Group 3 88.5％ 70.0％ About 503 yuan / g About 8.8L / g Group 4 89.4％ 65％ About 542 yuan / g About 9.5L / g Group 5 93.2％ 71.3％ About 494 yuan / g About 8.7L / g Group 6 92.9％ 68.3％ About 516 yuan / g About 9.1L / g Group 7 86.5％ 76％ About 463 yuan / g About 8.1L / g Group 8 98.5％ 60％ About 588 yuan / g About 10.3L / g Group 9 99.3％ 80.8％ About 441 yuan / g About 7.8L / g

[0076] From this comparative example, it can be seen that the purity and yield of Huperzine A (Group 9) of the present invention are higher than those of Groups 1 to 8 of the control, and it has the advantages of low production cost and less consumption of organic solvents.

[0077] Comparative Example 3

[0078] Huperzine A was obtained according to the method of Example 4 in Chinese Patent CN104262251B, specifically:

[0079] (1) Pretreatment: Take dried radix serrata medicinal materials (huperzine A content 0.053%), remove impurities, crush and pass through a 24-mesh sieve, and take 100 kg of the crushed medicinal materials for use;

[0080] (2) Ultrasonic countercurrent extraction: add 575 kg of water to the crushed medicinal materials and soak them twice, each time for 37 min at a soaking temperature of 43°C, and discard the soaking liquid; send the soaked medicinal materials into an ultrasonic countercurrent extraction device, add 1375 kg of alkaline ethanol with a volume fraction of 78% and a pH value of 9.04 for extraction, the ultrasonic frequency is 50 kHz, the extraction temperature is 56°C, the extraction time is 32 min, and after extraction, filter to obtain 1336 kg of extract;

[0081] (3) Membrane separation and purification: The extract obtained in step (2) is centrifuged using a butterfly centrifuge. After centrifugation, ultrafiltration is performed using an ultrafiltration membrane with a molecular weight cutoff of 3wD at an operating pressure of 0.5 to 1.0 bar; nanofiltration is then performed using a nanofiltration membrane with a molecular weight cutoff of 200D at an operating pressure of 5.0 to 15.0 bar to obtain a retentate;

[0082] (4) Concentration: The retentate obtained in step (3) was vacuum concentrated at a temperature of 56° C. and a vacuum degree of -0.08 to -0.04 MPa to obtain an extract, obtaining 7.73 kg of a paste;

[0083] (5) Acid dissolution: 88.5 kg of 0.75% by volume hydrochloric acid aqueous solution was added to the paste to dissolve it. After dissolution, the solution was filtered. Then, the pH of the filtrate was adjusted to 8.52 using 7.5% by mass sodium hydroxide solution to obtain 90.32 L of alkaline solution.

[0084] (6) Extraction: The alkaline solution obtained in step (5) was extracted with chloroform three times, each time for 27 min, using 340 L of chloroform each time. The chloroform phases were collected and combined to obtain 995.2 L of chloroform extracts from three extractions.

[0085] (7) Concentration: The 995.2 L chloroform extract obtained in step (6) was concentrated under vacuum at a temperature of 47° C. and a vacuum degree of -0.08 to -0.04 MPa until dryness to obtain 35.97 g of dry powder;

[0086] (8) Crystallization and recrystallization: 1780 mL of acetone solvent was added to dissolve the dry powder obtained in step (7), and the temperature was lowered to 5° C. and crystallized three times to obtain white crystals;

[0087] (9) Drying: The white crystals obtained in step (8) were vacuum dried at a temperature of 67° C. and a vacuum degree of -0.09 to -0.06 MPa to obtain 28.02 g of Huperzine A product.

[0088] The purity, yield, production cost and organic solvent consumption of this comparative example and Examples 1-3 of the present invention are shown in Table 3.

[0089] Table 3

[0090]

[0091]

[0092] As can be seen from Table 3, the yield of Huperzine A of the present invention is significantly higher than that of Comparative Example 3, and has the advantages of low production cost and low organic solvent consumption.

Claims

1. A method for extracting and purifying huperzine A, comprising the following steps: (1) Cut the Thousand Layer Pagoda into 2-3 cm segments and dry them. Before cutting the Thousand Layer Pagoda into 2-3cm segments, sort out the impurities and wash them with water; (2) adding alkaline ethanol to the dried Cassia bark in step (1), stirring and impregnating, then heating under stirring and refluxing for extraction, filtering, and concentrating to an ethanol concentration of 60%-65%, wherein the ratio of alkaline ethanol to dried Cassia bark is (7-10) mL:1 g; the alkaline ethanol concentration is 90-95%, and the pH value is 8.5-9.5; (3) stirring the concentrated solution in step (2), cooling it to below 10° C., refrigerating it, and filtering it; (4) concentrating the filtered solution in step (3) to an extract, adding an acidic aqueous solution to adjust the pH value to 2.8-3.0, filtering, adjusting the pH value of the filtered solution to 8.5-9.5 with an alkaline aqueous solution, and then drying the water to obtain a solid; (5) adding a mixed solvent of ethyl acetate and n-heptane to the solid matter in step (4), stirring at 60-70° C., then adding an alkaline aqueous solution with a pH value of 8.5-9.5 at 60-70° C. and stirring, keeping the temperature and standing to separate, discarding the aqueous solution, standing and cooling the obtained organic solution to 15-25° C. for crystallization, filtering, and drying to obtain huperzine A, wherein the volume ratio of ethyl acetate to n-heptane in the mixed solvent of ethyl acetate and n-heptane is 1:(2-3), and the mass ratio of the mixed solvent of ethyl acetate and n-heptane to the solid matter is (3-5):

1.

2. The method according to claim 1, characterized in that In the step (2), the stirring and soaking temperature is 40-45° C., and the stirring and soaking time is 2-4 hours; the heating and reflux extraction times are 3-5 times, and the extraction time each time is 2-4 hours.

3. The method according to claim 1, characterized in that In the step (3), the stirring temperature is room temperature, the stirring time is 8 to 15 minutes, and the standing refrigeration time is more than 24 hours.

4. The method according to claim 1, wherein In the step (4), the acidic aqueous solution is a hydrochloric acid solution or a sulfuric acid solution; and the alkaline aqueous solution is a sodium hydroxide solution, a potassium hydroxide solution or an ammonia solution.

5. The method according to claim 1, characterized in that In the step (5), the stirring time at 60-70° C. is 10-20 min.

6. The method according to claim 1, characterized in that In the step (5), the ratio of the alkaline aqueous solution with a pH value of 8.5 to 9.5 to the solid matter is (8-12) mL:1 g; the alkaline aqueous solution with a pH value of 8.5 to 9.5 is added at 60-70° C. and the stirring time is 20-40 min.

7. The method according to claim 1, characterized in that The number of extractions in step (5) is 3 to 4 times.

8. The method according to claim 1, characterized in that The crystallization time in step (5) is 1-2h.

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