Method for extracting flavone from astragalus membranaceus leaves

Abstract

The invention relates to a method for extracting flavone from astragalus membranaceus leaves, which comprises the following steps: mixing astragalus membranaceus leaf powder and water to obtain a first mixed material; sequentially carrying out first constant-temperature extraction treatment and first solid-liquid separation treatment on the first mixed material to obtain a first liquid-phase product and a first solid-phase product; the first solid-phase product and water are mixed to obtain a second mixed material, the second mixed material is sequentially subjected to second constant-temperature extraction treatment and second solid-liquid separation treatment to obtain a second liquid-phase product and a second solid-phase product, and the first liquid-phase product and the second liquid-phase product are mixed to obtain a liquid-phase product; mixing the liquid-phase product with anhydrous sodium sulfate to obtain a coarse impurity-removed liquid; and sequentially carrying out vacuum concentration treatment, first drying treatment and first crushing treatment on the coarse impurity-removed liquid to obtain a flavone powder product. According to the method, water is used as an extracting agent, constant-temperature extraction is combined with a secondary extraction process, and anhydrous sodium sulfate is used for adsorbing moisture and polar impurities in a liquid-phase product, so that the flavone in the astragalus membranaceus leaves can be efficiently extracted.

Description

Technical Field

[0001] This invention relates to the field of traditional Chinese medicine extraction technology, and in particular to a method for extracting flavonoids from Astragalus membranaceus leaves. Background Technology

[0002] Astragalus membranaceus, belonging to the genus Astragalus of the legume family, is a traditional and precious Chinese medicinal herb. Its roots, stems, and leaves are rich in flavonoids. As an important class of natural active ingredients, flavonoids possess various biological activities such as antioxidant, anti-inflammatory, immunomodulatory, and lipid-lowering effects, and have broad application prospects in the fields of medicine, food, and health products, with market demand continuously rising.

[0003] Currently, the development and utilization of Astragalus membranaceus mainly focuses on its roots, with a large amount of Astragalus membranaceus leaves being discarded as processing waste. This not only causes a serious waste of natural resources but also brings certain environmental pressures. Therefore, extracting flavonoids from Astragalus membranaceus leaves to achieve high-value utilization of Astragalus membranaceus leaf waste has become an important direction for the comprehensive development of Astragalus membranaceus resources, with significant economic value and environmental significance.

[0004] Among existing flavonoid extraction technologies from Astragalus membranaceus leaves, the traditional alcohol extraction method is widely used due to its simplicity and low cost. However, this process has significant technical drawbacks: the extraction parameters have not been systematically optimized, resulting in a low flavonoid extraction rate; improper temperature and time control during the extraction process can easily lead to oxidation and degradation of flavonoids, resulting in loss of biological activity; improper selection of concentration equipment leads to instability in the concentration process, which can easily cause product charring and a decrease in purity; at the same time, some processes suffer from high energy consumption, cumbersome operation, and difficulty in large-scale promotion, failing to meet the market demand for industrial production and high-quality flavonoid extracts.

[0005] In addition, the existing technology does not strictly control the variety of Astragalus leaf raw materials, resulting in inconsistent raw material quality, which in turn affects the flavonoid extraction effect and product quality stability. Some extraction processes do not have reasonable impurity removal and purification steps, or the impurity removal parameters are unreasonable, resulting in excessively high impurity content in the extract, which limits its application in high-end fields.

[0006] To address the shortcomings of the existing technologies, developing a new extraction method to extract flavonoids from Astragalus membranaceus leaves and improving the flavonoid extraction rate is an urgent problem to be solved. Summary of the Invention

[0007] To address the aforementioned technical problems, this invention provides a method for extracting flavonoids from Astragalus membranaceus leaves. Using water as the extraction solvent, a constant-temperature extraction combined with a secondary extraction process is employed. Anhydrous sodium sulfate is used to adsorb moisture and polar impurities in the liquid phase product. This effectively solves the technical pain points of existing processes, such as low extraction efficiency, poor retention of flavonoid activity, high energy consumption, and unstable product quality. It achieves efficient, green, and large-scale extraction of flavonoids from Astragalus membranaceus leaves, providing a practical technical solution for the comprehensive utilization of Astragalus membranaceus resources.

[0008] To achieve this objective, the present invention adopts the following technical solution: In a first aspect, the present invention provides a method for extracting flavonoids from Astragalus membranaceus leaves, the extraction method comprising the following steps: (1) The Astragalus leaf powder and water are mixed for the first time to obtain the first mixture; (2) The first mixture is subjected to a first constant temperature extraction treatment and a first solid-liquid separation treatment in sequence to obtain a first liquid phase product and a first solid phase product; (3) The first solid product and water are mixed for the second time to obtain a second mixture. The second mixture is subjected to a second constant temperature extraction treatment and a second solid-liquid separation treatment in sequence to obtain a second liquid product and a second solid product. The first liquid product and the second liquid product are mixed for the third time to obtain a liquid product. (4) The liquid product and anhydrous sodium sulfate are mixed for the fourth time to roughly remove impurities from the liquid product and obtain the roughly removed liquid. (5) The liquid after crude impurity removal is subjected to vacuum concentration, first drying and first pulverization in sequence to obtain flavonoid powder product.

[0009] This invention uses water as the extraction solvent to perform isothermal extraction of Astragalus membranaceus leaf powder. The solid phase product after the first isothermal extraction is then subjected to a second isothermal extraction to maximize the recovery of flavonoids from the raw material, ensuring thorough extraction, further improving the flavonoid extraction rate, and reducing waste of Astragalus membranaceus leaf resources. The first and second liquid phase products are then combined, and anhydrous sodium sulfate is used to fully adsorb trace amounts of water and some polar impurities from the liquid phase product. The anhydrous sodium sulfate precipitate is then removed, yielding a crude purified liquid. This crude purified liquid is then concentrated under reduced pressure and dried to obtain the flavonoid product. This invention uses water as the extraction solvent and does not add anhydrous ethanol or other organic solvents throughout the process, ensuring no organic solvent residue. Furthermore, this invention has advantages such as high extraction efficiency, low energy consumption, simple operation, and good preservation of flavonoid activity. It is suitable for small-scale laboratory preparation, industrial pilot-scale production, and large-scale industrial production. The vacuum concentration ring treatment uses a rotary evaporator to ensure a stable and controllable concentration process, further guaranteeing product quality.

[0010] It should be noted that this invention uses Astragalus leaves as raw material, which is different from the plant parts of Astragalus rhizome. Astragalus leaves have the characteristic of high efficiency in pretreatment. Therefore, flavonoids can be extracted efficiently from Astragalus leaves when water is used as the extraction agent, while Astragalus rhizome requires ethanol as the extraction agent.

[0011] As a preferred technical solution of the present invention, the preparation of the Astragalus leaf powder includes: washing the Astragalus leaves in sequence, drying them, pulverizing them, and sieving them to obtain the Astragalus leaf powder.

[0012] Preferably, the cleaning agent used in the cleaning process includes water.

[0013] Preferably, the temperature of the second drying process is 40~60°C, for example, it can be 40°C, 45°C, 50°C, 55°C or 60°C, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0014] Preferably, the sieving process includes: using a 60-120 mesh sieve for sieving, for example, it can be 60 mesh, 70 mesh, 80 mesh, 100 mesh or 120 mesh, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0015] In this invention, the astragalus leaves are first washed to remove mud and impurities. Then, a second drying process is performed to remove surface moisture. The dried astragalus leaves are then pulverized and passed through a 60-120 mesh sieve. The sieved powder is collected to obtain astragalus leaf powder with uniform particle size. In this invention, the astragalus leaf powder is subjected to constant temperature extraction, which can increase the contact area between the astragalus leaf powder and water and improve the flavonoid dissolution efficiency.

[0016] As a preferred technical solution of the present invention, the ratio of Astragalus leaf powder to water in the first mixture is 1g:(10~20)mL, for example, it can be 1g:10mL, 1g:12mL, 1g:15mL, 1g:18mL or 1g:20mL, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0017] Preferably, a first settling process is performed after the first mixing to obtain a first mixture.

[0018] Preferably, the first settling time is 20 to 40 minutes, for example, it can be 20 minutes, 25 minutes, 30 minutes, 35 minutes or 40 minutes, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0019] In this invention, a first settling treatment is performed after the first mixing, and the settling time is limited to ensure that the Astragalus leaf powder is fully soaked, so that water can penetrate into the Astragalus leaf powder and dissolve flavonoids.

[0020] As a preferred technical solution of the present invention, the temperature of the first constant temperature extraction treatment is 80~85℃, for example, it can be 80℃, 81℃, 82℃, 83℃, 84℃ or 85℃, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0021] This invention employs a constant-temperature extraction process. Compared to traditional extraction, the temperature control during this constant-temperature extraction process is more precise (temperature control accuracy is ±1℃), which balances flavonoid dissolution efficiency and activity retention. It avoids insufficient flavonoid dissolution due to excessively low temperatures, which would reduce the extraction rate. At the same time, it avoids flavonoid degradation and activity loss due to excessively high temperatures, and also prevents the aqueous solution from boiling over, ensuring operational safety.

[0022] Preferably, the extraction time of the first constant temperature extraction treatment is 180~210 min, for example, it can be 180 min, 190 min, 200 min or 210 min, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0023] This invention reduces energy consumption while ensuring flavonoid dissolution efficiency by limiting the extraction time of the first constant temperature extraction treatment. If the extraction time is too long, it will lead to increased energy consumption and excessive dissolution of impurities. If the extraction time is too short, the flavonoids will not be extracted sufficiently, affecting the extraction efficiency.

[0024] Preferably, the first constant temperature extraction process involves stirring every 20 to 30 minutes, for example, 20 minutes, 22 minutes, 24 minutes, 26 minutes, 28 minutes, or 30 minutes, but is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0025] Preferably, the stirring speed is 45~60 r / min, for example, it can be 45 r / min, 50 r / min, 55 r / min or 60 r / min, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0026] Preferably, the stirring time is ≥1 min, for example, it can be 1 min, 2 min, 3 min, 4 min or 5 min, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0027] This invention incorporates intermittent stirring during the constant-temperature extraction process to ensure sufficient contact between the Astragalus leaf powder and water, promoting the full dissolution of flavonoids into the aqueous phase system. Furthermore, compared to continuous stirring during constant-temperature extraction, intermittent stirring reduces energy consumption while maintaining extraction efficiency.

[0028] As a preferred technical solution of the present invention, the ratio of the first solid product to water in the second mixture is 1g:(10~15)mL, for example, it can be 1g:10mL, 1g:11mL, 1g:12mL, 1g:13mL, 1g:14mL or 1g:15mL, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0029] Preferably, the material-to-liquid ratio in the second mixture is the same as that in the first mixture.

[0030] Preferably, the temperature of the second constant temperature extraction treatment is 80~85℃, for example, it can be 80℃, 81℃, 82℃, 83℃, 84℃ or 85℃, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0031] Preferably, the extraction time of the second constant temperature extraction treatment is 150~180 min, for example, it can be 150 min, 160 min, 170 min or 180 min, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0032] This invention reduces energy consumption while ensuring flavonoid dissolution efficiency by limiting the extraction time of the second constant temperature extraction treatment. If the extraction time is too long, it will lead to increased energy consumption and excessive dissolution of impurities. If the extraction time is too short, the flavonoids will not be extracted sufficiently, affecting the extraction efficiency.

[0033] Preferably, the second constant temperature extraction process involves stirring every 20 to 30 minutes, for example, 20 minutes, 22 minutes, 24 minutes, 26 minutes, 28 minutes, or 30 minutes, but is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0034] In this invention, after the first and second constant temperature extraction treatments, the extract needs to be cooled to room temperature before the first and second solid-liquid separation treatments are performed respectively. This avoids flavonoid degradation caused by solid-liquid separation at high temperatures and reduces extract loss.

[0035] As a preferred embodiment of the present invention, the mass-to-volume ratio of anhydrous sodium sulfate to the liquid phase product in the fourth mixture is (0.005~0.01) g:1 mL, for example, it can be 0.005 g:1 mL, 0.006 g:1 mL, 0.007 g:1 mL, 0.008 g:1 mL, 0.009 g:1 mL or 0.01 g:1 mL, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0036] In this invention, during the coarse impurity removal process, the mass-to-volume ratio of anhydrous sodium sulfate to the liquid product is limited so that anhydrous sodium sulfate can adsorb water and polar impurities in the liquid product. If too much anhydrous sodium sulfate is used, it will lead to the loss of flavonoid adsorption. If too little anhydrous sodium sulfate is used, it will lead to incomplete impurity removal.

[0037] Preferably, the fourth mixing is carried out under stirring.

[0038] Preferably, the stirring time is 5 to 15 minutes, for example, 5 minutes, 7 minutes, 10 minutes, 12 minutes or 15 minutes, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0039] Preferably, after the fourth mixing, a second settling treatment and a third solid-liquid separation treatment are performed sequentially to obtain a coarsely purified liquid.

[0040] Preferably, the second settling time is 10 to 30 minutes, for example, 10 minutes, 15 minutes, 20 minutes, 25 minutes or 30 minutes, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0041] Preferably, the first solid-liquid separation process, the second solid-liquid separation process, and the third solid-liquid separation process each independently include a filtration process and / or a centrifugation process.

[0042] Preferably, the centrifugation speed is 7500~8500 r / min, for example, it can be 7500 r / min, 7800 r / min, 8000 r / min, 8200 r / min or 8500 r / min, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0043] Preferably, the centrifugation time is 10 to 20 minutes, for example, 10 minutes, 12 minutes, 14 minutes, 16 minutes, 18 minutes or 20 minutes, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0044] As a preferred embodiment of the present invention, the extraction method further includes purifying the crude impurity-removed liquid using a polyamide column before vacuum concentration, removing impurities by loading water onto the column after purification, and then eluting with water to obtain an eluent.

[0045] This invention purifies the crude impurity-removing solution to further improve the purity of the flavonoid extract, and then uses water to rinse the column after purification to remove water-soluble impurities.

[0046] Preferably, the polyamide column is activated before purification.

[0047] In this invention, the activation treatment involves immersing the polyamide column in deionized water for 20 to 24 hours, for example, 20 hours, 21 hours, 22 hours, 23 hours, or 24 hours, but is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0048] Preferably, the flow rate of the liquid after crude impurity removal in the purification process is 1.5~2.5 BV / h, for example, it can be 1.5 BV / h, 1.7 BV / h, 2.0 BV / h, 2.3 BV / h or 2.5 BV / h, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0049] Preferably, the water flow rate during the impurity removal process is 2~2.5 BV / h, for example, it can be 2 BV / h, 2.1 BV / h, 2.2 BV / h, 2.3 BV / h, 2.4 BV / h or 2.5 BV / h, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0050] Preferably, the water flow rate in the elution process is 1.5~2.5 BV / h, for example, it can be 1.5 BV / h, 1.8 BV / h, 2.0 BV / h, 2.2 BV / h or 2.5 BV / h, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0051] As a preferred technical solution of the present invention, the flavonoid concentrate is obtained after the vacuum concentration treatment.

[0052] Preferably, the temperature of the vacuum concentration process is 60~65℃, for example, it can be 60℃, 61℃, 62℃, 63℃, 64℃ or 65℃, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0053] Preferably, the vacuum degree of the reduced pressure concentration process is 0.06~0.08MPa, for example, it can be 0.06MPa, 0.065MPa, 0.07MPa, 0.075MPa or 0.08MPa, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0054] In this invention, temperature and vacuum level need to be strictly controlled during the vacuum concentration process, following the principle of "low temperature and slow concentration" to avoid product charring and flavonoid degradation caused by excessively rapid concentration; at the same time, the concentrate needs to be dried in time to prevent moisture absorption and clumping, which would affect product quality.

[0055] Preferably, the vacuum concentration process is accompanied by stirring.

[0056] Preferably, the density ratio of the flavonoid concentrate to the eluent is (1.2~1.25):1, for example, it can be 1.2:1, 1.21:1, 1.22:1, 1.23:1, 1.24:1 or 1.25:1, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0057] In this invention, continuous stirring is carried out during the vacuum concentration process to ensure uniform concentration and avoid local overheating that could lead to degradation or charring of flavonoids. Concentration is continued until the extract becomes viscous and the relative density reaches 1.20~1.25, at which point the concentration is stopped to obtain a flavonoid concentrate.

[0058] As a preferred technical solution of the present invention, the temperature of the first drying treatment is 50~60℃, for example, it can be 50℃, 52℃, 54℃, 56℃, 58℃ or 60℃, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0059] In this invention, by limiting the parameters of vacuum concentration and the first drying process, the activity of flavonoids is not lost and the water content of the flavonoid powder product meets the standard (≤8%).

[0060] Preferably, the average particle size of the flavonoid powder product is 150~200μm, for example, it can be 150μm, 160μm, 170μm, 180μm, 190μm or 200μm, but it is not limited to the listed values. Other unlisted values ​​within the above range are also applicable.

[0061] As a preferred technical solution of the present invention, the extraction method includes the following steps: (1) The leaves of Astragalus membranaceus are washed, dried at 40-60℃, and pulverized. They are then sieved using a 60-120 mesh screen to obtain Astragalus membranaceus leaf powder. The Astragalus membranaceus leaf powder and water are mixed in a material-liquid ratio of 1g:(10-20)mL and allowed to stand for 20-40min to obtain the first mixture. (2) The first mixture is subjected to a first constant temperature extraction treatment at a temperature of 80~85℃ and a time of 180~210min and a first solid-liquid separation treatment in sequence to obtain a first liquid phase product and a first solid phase product; (3) Mix the first solid product and water in a material-liquid ratio of 1g:(10~15)mL to obtain a second mixture. Then, subject the second mixture to a second constant temperature extraction treatment at a temperature of 80~85℃ and a time of 150~180min, and a second solid-liquid separation treatment to obtain a second liquid product and a second solid product. Finally, mix the first liquid product and the second liquid product in a third process to obtain a liquid product. (4) Anhydrous sodium sulfate and liquid phase product were mixed for the fourth time according to a mass-volume ratio of (0.005~0.01)g:1mL to remove impurities from the liquid phase product. After the fourth mixing, the second settling treatment and the third solid-liquid separation treatment were carried out for 10~30min in sequence to obtain the liquid after removing impurities. Then, the liquid after removing impurities was purified by using a polyamide column. After purification, water was used to remove impurities from the column. Then, water was used to elute the liquid to obtain the eluent. (5) The eluent was subjected to vacuum concentration at a temperature of 60~65℃ and a vacuum degree of 0.06~0.08MPa, followed by a first drying treatment and a first pulverizing treatment to obtain a flavonoid powder product with an average particle size of 150~200μm.

[0062] Compared with the prior art, the present invention has at least the following beneficial effects: This invention uses Astragalus membranaceus leaves as raw material and water as the extraction agent. It employs a constant temperature extraction combined with a secondary extraction process and uses anhydrous sodium sulfate to adsorb water and polar impurities in the liquid phase product. This achieves efficient, green, and large-scale extraction of flavonoids from Astragalus membranaceus leaves, with an extraction rate of over 12.1 mg / g. It realizes the resource utilization of waste, reduces environmental pollution, and practices the concept of environmental protection. Detailed Implementation

[0063] To facilitate understanding of the present invention, the following embodiments are provided. Those skilled in the art should understand that these embodiments are merely illustrative and should not be construed as limiting the scope of the invention.

[0064] The Astragalus leaves used in the following examples are from Guyang County, Baotou City, Inner Mongolia. Guyang County is located in the western section of the Daqing Mountains on the Inner Mongolia Plateau, with a climate suitable for Astragalus growth. It is one of the authentic and high-quality Astragalus producing areas, where the Astragalus produced is firm yet soft, powdery, rich in flavonoids and other nutrients, and of excellent quality, free from mold and insect infestation. The above limitations are only for clearly illustrating the technical solution of the present invention and are not considered as further limitations on the present invention.

[0065] Example 1 This embodiment provides a method for extracting flavonoids from Astragalus membranaceus leaves, the extraction method comprising the following steps: (1) First, wash the Astragalus leaves with water, then dry and pulverize the washed Astragalus leaves at a temperature of 50°C, and sieve them with an 80-mesh sieve to obtain Astragalus leaf powder. Mix the Astragalus leaf powder and water at a material-liquid ratio of 1g:15mL and let stand for 30 minutes to obtain the first mixture. (2) The first mixture is subjected to a first constant temperature extraction treatment at a temperature of 82°C for 200 min to obtain a first extract. The first extract is cooled to 25°C and then filtered to obtain a first liquid product and a first solid product. In the first constant temperature extraction treatment, a stirring treatment at a speed of 60 r / min for 3 min is performed every 20 min. (3) The first solid product and water are mixed in a second mixture at a ratio of 1g:10mL to obtain a second mixture. The second mixture is subjected to a second constant temperature extraction treatment at a temperature of 82℃ for 160min to obtain a second extract. The second extract is cooled to 25℃ and then filtered to obtain a second liquid product and a second solid product. The first liquid product and the second liquid product are mixed in a third mixture to obtain a liquid product. In the second constant temperature extraction treatment, a stirring treatment at a speed of 60r / min for 4min is performed every 20min. (4) Under stirring, anhydrous sodium sulfate and liquid phase product were mixed for 10 min at a mass-volume ratio of 0.007 g: 1 mL to remove impurities from the liquid phase product. After the fourth mixing, the product was subjected to a second settling treatment for 20 min and a centrifugation treatment at a speed of 8000 r / min for 15 min to obtain a liquid after removing impurities. The liquid after removing impurities was then purified using a polyamide column at a flow rate of 2 BV / h. After purification, the product was loaded onto the column with water at a flow rate of 2.5 BV / h to remove impurities. Then, the product was eluted with water at a flow rate of 2 BV / h to obtain an eluent. Before purification, the polyamide column was soaked in deionized water for 24 h to activate the polyamide column. (5) Under stirring, the eluent is concentrated under reduced pressure at a temperature of 63°C and a vacuum of 0.07MPa to obtain flavonoid concentrate; the density ratio of flavonoid concentrate to eluent is 1.22:1. Then, the flavonoid concentrate is subjected to a first drying treatment and a first pulverizing treatment at a temperature of 55°C to obtain flavonoid powder product with an average particle size of 180μm.

[0066] Example 2 This embodiment provides a method for extracting flavonoids from Astragalus membranaceus leaves, the extraction method comprising the following steps: (1) First, wash the Astragalus leaves with water, then dry and pulverize the washed Astragalus leaves at a temperature of 40°C, and sieve them with a 60-mesh sieve to obtain Astragalus leaf powder. Mix the Astragalus leaf powder and water at a material-liquid ratio of 1g:20mL and let stand for 40min to obtain the first mixture. (2) The first mixture is subjected to a first constant temperature extraction treatment at a temperature of 85°C for 180 min to obtain a first extract. The first extract is cooled to 25°C and then filtered to obtain a first liquid product and a first solid product. In the first constant temperature extraction treatment, a stirring treatment at a speed of 60 r / min for 5 min is performed every 30 min. (3) The first solid product and water are mixed in a second mixture at a ratio of 1g:15mL to obtain a second mixture. The second mixture is subjected to a second constant temperature extraction treatment at a temperature of 80℃ for 180min to obtain a second extract. The second extract is cooled to 25℃ and then centrifuged at a speed of 7500r / min for 20min to obtain a second liquid product and a second solid product. The first liquid product and the second liquid product are mixed in a third mixture to obtain a liquid product. In the second constant temperature extraction treatment, a stirring treatment at a speed of 60r / min for 3min is performed every 30min. (4) Under stirring, anhydrous sodium sulfate and liquid phase product were mixed for 15 min at a mass-volume ratio of 0.005 g: 1 mL to remove impurities from the liquid phase product. After the fourth mixing, the product was subjected to a second settling treatment for 30 min and a centrifugation treatment at a speed of 8500 r / min for 10 min to obtain a liquid after removing impurities. The liquid after removing impurities was then purified using a polyamide column at a flow rate of 1.5 BV / h. After purification, the product was loaded onto the column with water at a flow rate of 2.5 BV / h to remove impurities. Then, the product was eluted with water at a flow rate of 2.5 BV / h to obtain an eluent. Before purification, the polyamide column was soaked in deionized water for 22 h to activate the polyamide column. (5) Under stirring, the eluent is concentrated under reduced pressure at a temperature of 60°C and a vacuum of 0.08MPa to obtain flavonoid concentrate; the density ratio of flavonoid concentrate to eluent is 1.2:1. Then, the flavonoid concentrate is subjected to a first drying treatment and a first pulverizing treatment at a temperature of 60°C to obtain flavonoid powder product with an average particle size of 150μm.

[0067] Example 3 This embodiment provides a method for extracting flavonoids from Astragalus membranaceus leaves, the extraction method comprising the following steps: (1) First, the leaves of Astragalus membranaceus were washed with water. Then, the washed leaves of Astragalus membranaceus were dried and pulverized at a temperature of 60°C. The leaves were then sieved using a 120-mesh sieve to obtain powder of Astragalus membranaceus leaves. The powder of Astragalus membranaceus leaves and water were mixed in a ratio of 1g:10mL and allowed to stand for 20 minutes to obtain the first mixture. (2) The first mixture is subjected to a first constant temperature extraction treatment at a temperature of 80°C for 210 min to obtain a first extract. The first extract is cooled to 25°C and then filtered to obtain a first liquid product and a first solid product. In the first constant temperature extraction treatment, a stirring treatment at a speed of 60 r / min for 1 min is performed every 25 min. (3) The first solid product and water are mixed in a second mixture at a ratio of 1g:10mL to obtain a second mixture. The second mixture is subjected to a second constant temperature extraction treatment at a temperature of 85℃ for 150min to obtain a second extract. The second extract is cooled to 25℃ and then filtered to obtain a second liquid product and a second solid product. The first liquid product and the second liquid product are mixed in a third mixture to obtain a liquid product. In the second constant temperature extraction treatment, a stirring treatment at a speed of 60r / min for 3min is performed every 25min. (4) Under stirring, anhydrous sodium sulfate and liquid phase product were mixed for 5 min at a mass-volume ratio of 0.01 g: 1 mL to remove impurities from the liquid phase product. After the fourth mixing, the product was subjected to a second settling treatment for 10 min and a centrifugation treatment at a speed of 7500 r / min for 20 min to obtain a liquid after removing impurities. The liquid after removing impurities was then purified using a polyamide column at a flow rate of 2.5 BV / h. After purification, the product was loaded onto the column with water at a flow rate of 2.5 BV / h to remove impurities. Then, the product was eluted with water at a flow rate of 1.5 BV / h to obtain an eluent. Before purification, the polyamide column was soaked in deionized water for 20 h to activate the polyamide column. (5) Under stirring, the eluent is concentrated under reduced pressure at a temperature of 65°C and a vacuum of 0.06MPa to obtain flavonoid concentrate; the density ratio of flavonoid concentrate to eluent is 1.25:1. Then, the flavonoid concentrate is subjected to a first drying treatment and a first pulverizing treatment at a temperature of 50°C to obtain flavonoid powder product with an average particle size of 200μm.

[0068] Example 4 This embodiment provides a method for extracting flavonoids from Astragalus membranaceus leaves. The only difference from Embodiment 1 is that the time for the first constant temperature extraction in step (2) is adjusted from 200 min to 150 min. All other steps are the same as in Embodiment 1.

[0069] Example 5 This embodiment provides a method for extracting flavonoids from Astragalus membranaceus leaves. The only difference from Embodiment 1 is that the time for the first constant temperature extraction in step (2) is adjusted from 200 min to 250 min. All other steps are the same as in Embodiment 1.

[0070] Example 6 This embodiment provides a method for extracting flavonoids from Astragalus membranaceus leaves. The only difference from Embodiment 1 is that the time for the second constant temperature extraction in step (3) is adjusted from 160 min to 120 min. All other steps are the same as in Embodiment 1.

[0071] Example 7 This embodiment provides a method for extracting flavonoids from Astragalus membranaceus leaves. The only difference from Example 1 is that the time for the second constant temperature extraction in step (3) is adjusted from 160 min to 220 min. All other steps are the same as in Example 1.

[0072] Example 8 This embodiment provides a method for extracting flavonoids from Astragalus membranaceus leaves. The only difference from Example 1 is that, except that the first constant temperature extraction is followed by direct filtration without cooling, the method is the same as Example 1.

[0073] Example 9 This embodiment provides a method for extracting flavonoids from Astragalus membranaceus leaves. The only difference from Example 1 is that the mass-volume ratio of anhydrous sodium sulfate and liquid phase product in the fourth mixing step (4) is adjusted from 0.007g:1mL to 0.001g:1mL. All other aspects are the same as in Example 1.

[0074] Example 10 This embodiment provides a method for extracting flavonoids from Astragalus membranaceus leaves. The only difference from Example 1 is that the mass-volume ratio of anhydrous sodium sulfate and liquid phase product in the fourth mixing step (4) is adjusted from 0.007 g: 1 mL to 0.02 g: 1 mL. All other steps are the same as in Example 1.

[0075] Example 11 This embodiment provides a method for extracting flavonoids from Astragalus membranaceus leaves. The only difference from Example 1 is that the temperature of the vacuum concentration process is adjusted from 63°C to 70°C. All other aspects are the same as in Example 1.

[0076] Comparative Example 1 This comparative example provides a method for extracting flavonoids from Astragalus membranaceus leaves. The only difference from Example 1 is that, except that the first constant temperature extraction treatment at 82°C in step (2) is changed to a first extraction treatment at 82°C, the rest are the same as in Example 1.

[0077] In this comparative example, the extraction process was affected by temperature fluctuations, which led to a decrease in the bioactivity of flavonoids. However, constant temperature extraction can improve the quality of flavonoids.

[0078] Comparative Example 2 This comparative example provides a method for extracting flavonoids from Astragalus membranaceus leaves. The only difference from Example 1 is that, except that the second constant temperature extraction treatment at 82°C in step (3) is changed to a second extraction treatment at 82°C, the rest are the same as in Example 1.

[0079] In this comparative example, the extraction process was affected by temperature fluctuations, which led to a decrease in the bioactivity of flavonoids. However, constant temperature extraction can improve the quality of flavonoids.

[0080] Comparative Example 3 This comparative example provides a method for extracting flavonoids from Astragalus membranaceus leaves. The only difference from Example 1 is that step (4) does not include crude impurity removal, i.e., the liquid phase product is directly purified. All other steps are the same as in Example 1.

[0081] The masses of Astragalus leaf powder and flavonoid powder products were weighed separately, and the flavonoid extraction rate was calculated using the formula: Flavonoid extraction rate (mg / g) = Mass of flavonoid powder product (mg) / Astragalus leaf powder (g). The bioactivity of the flavonoid powder product was then tested using a UV spectrophotometer. A flavonoid bioactivity IC50 of ≤200 μg / mL was considered good, IC50 >200 μg / mL and ≤300 μg / mL was considered fair, and IC50 >300 μg / mL was considered poor. The flavonoid powder product was dried at 100℃ to constant weight, and the results were then analyzed using the formula: , The water content of the flavonoid powder product was calculated, and the test results are shown in Table 1.

[0082] Table 1 The test results show that: (1) As can be seen from Examples 1 to 3, the present invention uses Astragalus leaves as raw material, water as extractant, constant temperature extraction combined with secondary extraction process, and anhydrous sodium sulfate adsorbs water and polar impurities in liquid phase product, thereby achieving efficient, green and large-scale extraction of flavonoids from Astragalus leaves, and the flavonoid extraction rate can reach more than 12.1 mg / g.

[0083] (2) As can be seen from Examples 1 and 4-5, the first constant temperature extraction time in Example 1 was 200 min, and the flavonoid extraction rate was 13.6 mg / g, with good flavonoid bioactivity. In Example 4, the first constant temperature extraction time was 150 min, and the flavonoid extraction rate was 12.5 mg / g, with good flavonoid bioactivity. In Example 5, the first constant temperature extraction time was 250 min, and the flavonoid extraction rate was 13.4 mg / g, with poor flavonoid bioactivity. This shows that by specifically limiting the extraction time of the first constant temperature extraction, the present invention can reduce energy consumption while ensuring flavonoid dissolution efficiency. If the extraction time is too short, the flavonoid extraction will be insufficient, affecting the extraction efficiency. If the extraction time is too long, it will lead to increased energy consumption, excessive dissolution of impurities, and affect the flavonoid bioactivity.

[0084] (3) As can be seen from Examples 1 and 6-7, the second constant temperature extraction time in Example 1 was 160 min, and the flavonoid extraction rate was 13.6 mg / g, with good flavonoid bioactivity. In Example 6, the second constant temperature extraction time was 120 min, and the flavonoid extraction rate was 12.4 mg / g, with good flavonoid bioactivity. In Example 7, the second constant temperature extraction time was 220 min, and the flavonoid extraction rate was 13.2 mg / g, with average flavonoid bioactivity. This shows that by specifically limiting the extraction time of the second constant temperature extraction, the present invention can reduce energy consumption while ensuring flavonoid dissolution efficiency. If the extraction time is too short, the flavonoid extraction will be insufficient, affecting the extraction efficiency. If the extraction time is too long, it will lead to increased energy consumption, excessive dissolution of impurities, and affect the flavonoid bioactivity.

[0085] (4) As can be seen from Examples 1 and 8, after the first and second constant temperature extraction treatments, the extract is cooled to room temperature before the first and second solid-liquid separation treatments are performed respectively. This can avoid the degradation of flavonoids caused by solid-liquid separation at high temperature and improve the flavonoid extraction rate.

[0086] (5) As can be seen from Examples 1 and 9-10, in Example 1, the mass-volume ratio of anhydrous sodium sulfate to liquid product in the fourth mixture is 0.007 g: 1 mL, and the flavonoid extraction rate is 13.6 mg / g. In Example 9, the mass-volume ratio of anhydrous sodium sulfate to liquid product in the fourth mixture is 0.001 g: 1 mL, and the flavonoid extraction rate is 12.7 mg / g. In Example 10, the mass-volume ratio of anhydrous sodium sulfate to liquid product in the fourth mixture is 0.02 g: 1 mL, and the flavonoid extraction rate is 12.9 mg / g. This shows that the present invention limits the mass-volume ratio of anhydrous sodium sulfate to liquid product during the crude impurity removal process, so that anhydrous sodium sulfate can adsorb water and polar impurities in the liquid product. If the amount of anhydrous sodium sulfate is too small, the impurity removal will be incomplete. If the amount of anhydrous sodium sulfate is too large, the flavonoid adsorption will be lost, reducing the flavonoid extraction rate.

[0087] (6) As can be seen from Examples 1 and 11, by limiting the temperature during the vacuum concentration process, the present invention follows the principle of "low temperature and slow concentration", which can avoid the product charring, flavonoid degradation and reduced flavonoid bioactivity caused by excessively fast concentration, and improve the flavonoid extraction rate.

[0088] (7) As can be seen from Example 1 and Comparative Examples 1-2, the present invention uses constant temperature extraction treatment to control the temperature within the range of 82±1℃, which can take into account both the flavonoid dissolution efficiency and activity retention. However, if extraction treatment is used, the temperature fluctuation will lead to a serious reduction in the biological activity of flavonoids and affect the flavonoid extraction rate.

[0089] (8) As can be seen from Example 1 and Comparative Example 3, the present invention can significantly improve the flavonoid extraction rate by using anhydrous sodium sulfate to adsorb water and polar impurities in the liquid phase product.

[0090] In summary, this invention uses Astragalus membranaceus leaves as raw material and water as the extraction agent. It employs a constant temperature extraction combined with a secondary extraction process and uses anhydrous sodium sulfate to adsorb water and polar impurities in the liquid phase product. This achieves efficient, green, and large-scale extraction of flavonoids from Astragalus membranaceus leaves, with an extraction rate of over 12.1 mg / g. This realizes the resource utilization of waste, reduces environmental pollution, and practices the concept of environmental protection.

[0091] The applicant declares that the above description is only a specific embodiment of the present invention, but the protection scope of the present invention is not limited thereto. Those skilled in the art should understand that any changes or substitutions that can be easily conceived by those skilled in the art within the technical scope disclosed in the present invention fall within the protection and disclosure scope of the present invention.

Claims

1. A method for extracting flavonoids from Astragalus membranaceus leaves, characterized in that, The extraction method includes the following steps: (1) The Astragalus leaf powder and water are mixed for the first time to obtain the first mixture; (2) The first mixture is subjected to a first constant temperature extraction treatment and a first solid-liquid separation treatment in sequence to obtain a first liquid phase product and a first solid phase product; (3) The first solid product and water are mixed for the second time to obtain a second mixture. The second mixture is subjected to a second constant temperature extraction treatment and a second solid-liquid separation treatment in sequence to obtain a second liquid product and a second solid product. The first liquid product and the second liquid product are mixed for the third time to obtain a liquid product. (4) The liquid product and anhydrous sodium sulfate are mixed for the fourth time to roughly remove impurities from the liquid product and obtain the roughly removed liquid. (5) The liquid after crude impurity removal is subjected to vacuum concentration, first drying and first pulverization in sequence to obtain flavonoid powder product.

2. The extraction method according to claim 1, characterized in that, The preparation of the Astragalus leaf powder includes: sequentially washing the Astragalus leaves, drying them, pulverizing them, and sieving them to obtain the Astragalus leaf powder; Preferably, the cleaning agent used in the cleaning process includes water; Preferably, the temperature of the second drying treatment is 40~60℃; Preferably, the screening process includes screening using a 60-120 mesh screen.

3. The extraction method according to claim 1 or 2, characterized in that, In the first mixture, the ratio of Astragalus leaf powder to water is 1g:(10~20)mL; Preferably, after the first mixing, a first settling treatment is performed to obtain a first mixture. Preferably, the first settling time is 20-40 minutes.

4. The extraction method according to any one of claims 1-3, characterized in that, The temperature of the first constant temperature extraction treatment is 80~85℃; Preferably, the extraction time for the first constant temperature extraction treatment is 180~210 min; Preferably, the stirring process is performed every 20-30 minutes during the first constant temperature extraction treatment; Preferably, the stirring time is ≥1 min.

5. The extraction method according to claim 3, characterized in that, In the second mixture, the ratio of the first solid product to water is 1 g:(10~15) mL; Preferably, the material-to-liquid ratio in the second mixture is the same as that in the first mixture; Preferably, the temperature of the second constant temperature extraction treatment is 80~85℃; Preferably, the extraction time for the second constant temperature extraction treatment is 150-180 min; Preferably, the second constant temperature extraction process involves stirring every 20 to 30 minutes.

6. The extraction method according to any one of claims 1-5, characterized in that, The mass-to-volume ratio of anhydrous sodium sulfate to the liquid phase product in the fourth mixture is (0.005~0.01) g: 1 mL; Preferably, the fourth mixing is carried out under stirring; Preferably, the stirring time is 5-15 minutes; Preferably, after the fourth mixing, a second settling treatment and a third solid-liquid separation treatment are performed sequentially to obtain a coarsely purified liquid; Preferably, the second settling time is 10-30 minutes.

7. The extraction method according to any one of claims 1-6, characterized in that, The extraction method further includes purifying the crude impurity-removed liquid using a polyamide column before vacuum concentration, removing impurities by loading water onto the column after purification, and then eluting with water to obtain the eluent. Preferably, the polyamide column is activated before purification. Preferably, the flow rate of the liquid after crude impurity removal in the purification process is 1.5~2.5 BV / h; Preferably, the water flow rate during the impurity removal process on the upper column is 2~2.5 BV / h; Preferably, the water flow rate in the elution process is 1.5~2.5 BV / h.

8. The extraction method according to claim 7, characterized in that, The flavonoid concentrate was obtained after the vacuum concentration treatment; Preferably, the temperature of the vacuum concentration treatment is 60~65℃; Preferably, the vacuum degree of the reduced pressure concentration process is 0.06~0.08 MPa; Preferably, the vacuum concentration process is accompanied by stirring; Preferably, the density ratio of the flavonoid concentrate to the eluent is (1.2~1.25):

1.

9. The extraction method according to any one of claims 1-8, characterized in that, The temperature of the first drying treatment is 50~60℃; Preferably, the average particle size of the flavonoid powder product is 150~200μm.

10. The extraction method according to any one of claims 1-9, characterized in that, The extraction method includes the following steps: (1) The leaves of Astragalus membranaceus are washed, dried at 40-60℃, and pulverized. They are then sieved using a 60-120 mesh screen to obtain Astragalus membranaceus leaf powder. The Astragalus membranaceus leaf powder and water are mixed in a material-liquid ratio of 1g:(10-20)mL and allowed to stand for 20-40min to obtain the first mixture. (2) The first mixture is subjected to a first constant temperature extraction treatment at a temperature of 80~85℃ and a time of 180~210min and a first solid-liquid separation treatment in sequence to obtain a first liquid phase product and a first solid phase product; (3) Mix the first solid product and water in a material-liquid ratio of 1g:(10~15)mL to obtain a second mixture. Then, subject the second mixture to a second constant temperature extraction treatment at a temperature of 80~85℃ and a time of 150~180min, and a second solid-liquid separation treatment to obtain a second liquid product and a second solid product. Finally, mix the first liquid product and the second liquid product in a third process to obtain a liquid product. (4) Anhydrous sodium sulfate and liquid phase product were mixed for the fourth time according to a mass-volume ratio of (0.005~0.01)g:1mL to remove impurities from the liquid phase product. After the fourth mixing, the second settling treatment and the third solid-liquid separation treatment were carried out for 10~30min in sequence to obtain the liquid after removing impurities. Then, the liquid after removing impurities was purified by using a polyamide column. After purification, water was used to remove impurities from the column. Then, water was used to elute the liquid to obtain the eluent. (5) The eluent was subjected to vacuum concentration at a temperature of 60~65℃ and a vacuum degree of 0.06~0.08MPa, followed by a first drying treatment and a first pulverizing treatment to obtain a flavonoid powder product with an average particle size of 150~200μm.

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