Method for extracting berberine from coptis chinensis based on deep-eutectic solvent

Abstract

The invention relates to the field of plant extraction, and particularly discloses a method for extracting berberine from coptis chinensis based on a deep-eutectic solvent, and the method comprises the following steps: S1, drying coptis chinensis, and crushing to obtain coptis chinensis powder; s2, preparing a deep eutectic solvent from choline chloride, ethylene glycol and polyethylene glycol 400 according to a molar ratio of 1: (1.8-2.2): (0.8-1.2); s3, adding the coptis chinensis powder into the eutectic solvent, performing ultrasonic-assisted extraction at 45-50 DEG C for 30-40 minutes, and centrifuging to obtain supernate and filter residues; and S4, repeating the operation in the step S3 on the filter residues twice, merging the supernate, and then performing post-treatment on the supernate to obtain berberine. The method has the characteristic that berberine is efficiently extracted through the eutectic solvent by taking coptis chinensis as a raw material.

Description

Technical Field

[0001] This application relates to the field of plant extraction, and more specifically, it relates to a method for extracting berberine from Coptis chinensis using a eutectic solvent. Background Technology

[0002] Berberine, the core active ingredient of the traditional Chinese medicine Coptis chinensis, has a wide range of pharmacological effects, including antibacterial, anti-inflammatory, and hypoglycemic properties, and its demand continues to grow in the fields of pharmaceuticals, health products, and food additives.

[0003] However, current berberine extraction mainly employs solvent extraction, primarily using organic solvents such as ethanol and methanol. This requires high-temperature reflux extraction for 6-8 hours, resulting in high energy consumption and potential thermal degradation of the quaternary ammonium base structure of berberine, leading to low extraction rates and insufficient solvent recovery. Water-based extraction is also used, but berberine's solubility in water is only 0.003 g / mL (25℃), requiring pH adjustment to 1-2 to form hydrochloride salts to improve solubility, complicating subsequent desalting steps. While novel solvents like ionic liquids offer strong solubility, they are expensive, and some ionic liquids exhibit irreversible adsorption of alkaloids, further reducing actual extraction rates. Traditional countercurrent water extraction requires 8-10 stages of extraction towers, resulting in large equipment footprints. Furthermore, berberine easily forms micelles in the continuous phase, causing mass transfer efficiency to decrease over time, with a significant drop in extraction rate after 4 hours, ultimately impacting overall extraction efficiency.

[0004] Given the various problems with current traditional methods for berberine extraction, new extraction methods are needed. Eutectic solvents (DESs), as third-generation green solvents, can achieve precise design of polarity ranges through the control of the molar ratio of hydrogen bond acceptors and donors. They have broad application prospects in the field of plant extraction. Literature records the extraction of berberine hydrochloride using a eutectic solvent solution composed of carboxylic acids and polyols from Phellodendron amurense. Compared to Phellodendron amurense, Coptis chinensis contains a higher berberine content. Therefore, using Coptis chinensis as a raw material with eutectic solvents is of great significance for achieving large-scale berberine extraction. Summary of the Invention

[0005] In order to achieve high-efficiency extraction of berberine from Coptis chinensis using a eutectic solvent, this application provides a method for extracting berberine from Coptis chinensis using a eutectic solvent.

[0006] This application provides a method for extracting berberine from Coptis chinensis using a eutectic solvent, employing the following technical solution: A method for extracting berberine from Coptis chinensis using a eutectic solvent includes the following steps: S1. After drying, Coptis chinensis is pulverized to obtain Coptis chinensis powder; S2. A eutectic solvent was prepared by mixing choline chloride, ethylene glycol, and polyethylene glycol 400 in a molar ratio of 1:(1.8-2.2):(0.8-1.2). S3. Add Coptis chinensis powder to a eutectic solvent and extract with ultrasonic assistance at 45-50℃ for 30-40 min. After centrifugation, obtain the supernatant and filter residue. S4. Repeat step S3 with the filter residue twice, combine the supernatants, and then post-process the supernatant to obtain berberine.

[0007] By adopting the above technical solution, this application uses choline chloride as a hydrogen bond acceptor, providing a positive charge center to form ionic interactions with the quaternary ammonium base structure of berberine, thereby enhancing solubility. Ethylene glycol, as a short-chain hydrogen bond donor, lowers the melting point of the system while penetrating the plant cell wall, promoting the release of berberine. Polyethylene glycol 400, as a long-chain regulator, encapsulates berberine molecules through hydrophobic interactions, reducing its non-specific binding with other impurities. Compared to the current binary system of choline chloride and ethylene glycol, which is prone to decreased solubility due to hydrogen bond saturation, this application introduces polyethylene glycol 400 to regulate solvent polarity, achieving highly selective extraction of berberine. Finally, this application uses choline chloride-ethylene glycol-polyethylene glycol 400 as a ternary eutectic solvent to enhance the solubility of berberine while reducing co-extraction of other non-target components in Coptis chinensis, thus improving selectivity.

[0008] Based on this, this application uses low-temperature extraction to avoid the heat-sensitive degradation of berberine, followed by ultrasonic-assisted treatment to utilize the cavitation effect to disrupt the cell wall, shorten the extraction time, and facilitate the dissolution of berberine. Finally, the synergistic treatment of low-melting solvent at low temperature and ultrasonic treatment solves the problem of low extraction efficiency of traditional berberine.

[0009] Optionally, in step S3, after mixing the eutectic solvent and the micro / nano bubble water, Coptis chinensis powder is added, and the amount of micro / nano bubble water added is 5-8% of the volume of the eutectic solvent. The particle size of the micro / nano bubbles in the micro / nano bubble water is 200 nm-10 μm, and the bubble concentration in the micro / nano bubble water is 10. 6 -10 7 per mL.

[0010] By adopting the above technical solution, since the viscosity of eutectic solvents is usually higher than that of traditional solvents, it will affect the mass transfer efficiency, thereby affecting the dissolution and extraction effect of berberine. Therefore, in this application, when extracting with eutectic solvents, the eutectic solvent is mixed with micro-nano bubble water and then added to Coptis chinensis powder for extraction. The addition of water reduces the hydrogen bond network in the eutectic solvent, reduces the viscosity, and helps it to quickly penetrate into the interior of Coptis chinensis cells, promoting the release and diffusion of berberine. Moreover, the micro-nano bubbles continuously break during the rising process in the water, generating local micro-jet and shock waves, which enhances the penetration of the solvent into Coptis chinensis cells and the release of berberine. It can also enhance the ultrasonic cavitation effect, further improving the extraction efficiency of berberine.

[0011] Optionally, in step S3, the ratio of Coptis chinensis powder to eutectic solvent is 20-25 mL / g.

[0012] Optionally, the eutectic solvent in step S2 is prepared by the following method: After stirring choline chloride and ethylene glycol at 80-90℃ for 2-2.5h, the mixture is cooled to 50-55℃, polyethylene glycol 400 is added, and stirring is continued for 20-30min to obtain a eutectic solvent.

[0013] By adopting the above technical solution, choline chloride and ethylene glycol are first stirred at high temperature and then cooled before adding polyethylene glycol 400, which avoids the degradation of polyethylene glycol 400 due to high temperature and ensures the uniformity of the eutectic solvent in the ternary system.

[0014] Optionally, the specific operation for supernatant post-treatment in step S4 is as follows: S4-1. First, adjust the pH of the combined supernatant to 2-3; S4-2. Then add ammonium sulfate to the supernatant until saturated, and then shake at low temperature for 10-20 minutes. S4-3. Add a mixed solvent of ethyl acetate and n-hexane, let stand at low temperature for 30-40 min, and then centrifuge to obtain organic phase A, which is mainly composed of berberine, and aqueous phase B. S4-4. After washing organic phase A with water, the organic solvent is rotary evaporated to obtain crude extract of Coptis chinensis, which is then purified to obtain berberine; the aqueous phase B is distilled under reduced pressure to recover the eutectic solvent.

[0015] By adopting the above technical solution, in order to recover berberine and eutectic solvent in this application, the pH value of the supernatant is first adjusted to 2-3. The hydrogen ion concentration in the system increases, berberine is converted into free base, the polarity decreases, and it is more soluble in the organic phase. Moreover, the acidification treatment will reduce the viscosity of the eutectic solvent due to the partial destruction of the hydrogen bond network by hydrogen ions, which is beneficial to the subsequent countercurrent extraction treatment. Then, ammonium sulfate is added. As a strong electrolyte, ammonium sulfate produces a large number of ammonia ions and sulfate ions upon dissolution. Through salting out, the activity coefficient of berberine in the aqueous phase decreases, forcing it to transfer to the organic phase. Moreover, the salting out effect is weak in eutectic solvents such as ethylene glycol, reducing co-extraction. Low-temperature shaking treatment is then performed to reduce the viscosity of the eutectic solvent and accelerate the separation of the two phases. Then, a mixed solvent of ethyl acetate and n-hexane is added. Ethyl acetate has limited solubility for moderately polar berberine. Combined with n-hexane, the polarity of the system can be reduced, enhancing the solubility of berberine in the mixed solvent, making the polarities of the two more matched, thus achieving the extraction of berberine. In addition, the addition of n-hexane can reduce the miscibility of ethyl acetate and strengthen the separation of the two phases. Finally, the organic phase is rotary evaporated to obtain crude berberine. The crude berberine is further recrystallized for purification to obtain berberine. Ethyl glycol and polyethylene glycol 400 are recovered by vacuum distillation in the aqueous phase.

[0016] Optionally, in step S4-3, the volume ratio of ethyl acetate to n-hexane in the mixed solvent is 7:(2.5-3.5).

[0017] By adopting the above technical solution and controlling the ratio of ethyl acetate to n-hexane, the solubility of berberine is improved, the polarity of the mixed solvent is more similar to that of berberine, and the extraction of berberine is achieved through the principle of "like dissolves like".

[0018] Optionally, the aqueous phase B obtained in step S4-3 is further subjected to aqueous two-phase extraction and countercurrent partitioning. The specific operations are as follows: Aqueous two-phase extraction: The pH value of aqueous phase B is adjusted to 2-3, and then a mixed aqueous solution of PEG-6000 and ammonium sulfate is added. After shaking and mixing, the mixture is centrifuged to obtain an upper phase containing berberine and PEG and a lower phase containing eutectic solvent and ammonium sulfate. Countercurrent partitioning: Ethyl acetate and n-hexane are mixed at a volume ratio of 1:(1-1.1) as the mobile phase and countercurrently extracted with the upper phase obtained from the aqueous two-phase extraction to obtain an organic phase C containing berberine and an aqueous phase D. Then, step S4-4 is performed, and the organic phase C and organic phase A are combined, washed with water, and rotary evaporated to obtain crude berberine extract. The aqueous phase D is distilled under reduced pressure to recover the eutectic solvent.

[0019] By adopting the above technical solution, when the supernatant is subjected to only single-stage extraction in this application, a small amount of berberine will remain in aqueous phase B, leading to the recovery of berberine. Therefore, this application also performs two-phase extraction and multi-stage partitioning on aqueous phase B. First, when PEG and ammonium sulfate are mixed, the hydroxyl groups of PEG form hydrogen bonds with water, while the ions of ammonium sulfate compete for water molecules through water-water interaction. Then, the PEG molecular chains encapsulate berberine through hydrophobic interactions, forming a micelle structure, which increases its apparent solubility. Under acidic conditions, berberine exists in molecular form, with enhanced hydrophobicity, and is easily soluble in the PEG phase, resulting in the separation of the system into an upper phase rich in PEG and berberine and a lower phase rich in salt, achieving the separation of berberine from the eutectic solvent and impurities. Then, the upper phase and the mobile phase are subjected to multi-stage countercurrent partitioning to transfer berberine to the mobile phase, thereby achieving high-purity extraction of berberine and improving both the extraction rate and purity of berberine.

[0020] Optionally, during aqueous two-phase extraction, the mass concentration of PEG-6000 in the mixed aqueous solution is 20-25%, and the mass concentration of ammonium sulfate is 15-20%.

[0021] Optionally, in step S4-4, the aqueous phase D is first evaporated and crystallized to precipitate supersaturated ammonium sulfate crystals, filtered, and then the filtrate is passed through an ultrafiltration membrane with a molecular weight cutoff of 6000 Da. Finally, it is distilled under reduced pressure at -0.095 MPa and 80-100 °C to obtain the recovered eutectic solvent.

[0022] Optionally, in step S4-4, the parameters for the rotary evaporation of organic phase C and organic phase A after water washing are: rotary evaporation under reduced pressure conditions of 40-50℃ and -0.09-(-0.1)MPa.

[0023] In summary, this application has the following beneficial effects: 1. This application introduces polyethylene glycol 400 to adjust solvent polarity, using choline chloride-ethylene glycol-polyethylene glycol 400 as a ternary eutectic solvent to achieve highly selective extraction of berberine, enhance the solubility of berberine, and reduce co-extraction of other non-target components in Coptis chinensis, thereby improving selectivity. Based on this, this application uses low-temperature extraction to avoid the heat-sensitive degradation of berberine, and then uses ultrasonic-assisted treatment to utilize the cavitation effect to destroy the cell wall, shorten the extraction time, and facilitate the dissolution of berberine. Finally, through the synergistic treatment of the eutectic solvent at low temperature and ultrasonic treatment, the problem of low extraction efficiency of traditional berberine is solved.

[0024] 2. In this application, during the extraction with a eutectic solvent, the eutectic solvent is mixed with micro-nano bubble water before adding Coptis chinensis powder for extraction. The addition of water reduces the hydrogen bond network in the eutectic solvent, lowers the viscosity, and helps it to quickly penetrate into the Coptis chinensis cells, promoting the release and diffusion of berberine. Moreover, the micro-nano bubbles continuously break during the rising process in the water, generating local micro-jet streams and shock waves, which enhances the penetration of the solvent into the Coptis chinensis cells and the release of berberine. Furthermore, it can enhance the ultrasonic cavitation effect, further improving the extraction efficiency of berberine. 3. In this application, after the supernatant is extracted with a eutectic solvent, it is first subjected to single-stage extraction, and then the aqueous phase is subjected to two-phase extraction and countercurrent partitioning. Berberine in the aqueous phase is further collected and purified, and finally berberine with higher extraction rate and higher purity is obtained. Moreover, the eutectic solvent is recovered and can be recycled. Detailed Implementation

[0025] The following detailed description of this application is provided in conjunction with the embodiments. It should be noted that: unless otherwise specified, the conditions in the following embodiments are performed under conventional conditions or conditions recommended by the manufacturer. Unless otherwise specified, the raw materials used in the following embodiments are all from commercially available sources.

[0026] The low-temperature conditions in the following examples are circulating cooling water conditions, which are carried out at a low temperature of 5°C.

[0027] Example 1 A method for extracting berberine from Coptis chinensis using a eutectic solvent includes the following steps: S1. Dry Coptis chinensis until the moisture content is ≤5%, then pulverize it and sieve it to obtain Coptis chinensis powder with a mesh size of 60-80. S2. After heating choline chloride and ethylene glycol in an oil bath and stirring at 85°C for 2 hours, cool to 50°C and add polyethylene glycol 400. Continue stirring for 25 minutes to obtain a eutectic solvent. The molar ratio of choline chloride, ethylene glycol and polyethylene glycol 400 is 1:2:1. S3. Mix the eutectic solvent with water and add 1 kg of Coptis chinensis powder. The amount of water added is 6% of the volume of the eutectic solvent. The ratio of Coptis chinensis powder to eutectic solvent is 22 mL / g (that is, 1 g of Coptis chinensis powder is added to 22 mL of eutectic solvent). Extract with pulsed ultrasound at 45℃ for 35 min (ultrasound frequency is 40 kHz, power is 200 W, turn on for 2 seconds and turn off for 1 second). Then, centrifuge at 5000 rpm to obtain the supernatant and filter residue. S4. Repeat step S3 with the filter residue twice, combine the supernatants, and then post-process the supernatant to obtain berberine. The specific post-processing steps are as follows: S4-1. First, adjust the pH of the combined supernatant to 2.5; S4-2. Then add ammonium sulfate to the supernatant until saturated, and then shake at low temperature for 15 minutes. S4-3, then add ethyl acetate and n-hexane in a volume ratio of 7:3 to prepare a mixed solvent, let it stand at low temperature for 35 min and then centrifuge to obtain organic phase A, mainly composed of berberine and aqueous phase B. S4-4. After washing organic phase A with water, the organic solvent was rotary evaporated at 45℃ and -0.09MPa to obtain crude berberine extract. Then, the crude berberine extract was thermally dissolved in hot ethanol at 70℃, filtered, cooled and crystallized, and dried to obtain berberine extract. The eutectic solvent was recovered by vacuum distillation of aqueous phase B at -0.09 MPa and 90 °C.

[0028] Example 2 A method for extracting berberine from Coptis chinensis using a eutectic solvent includes the following steps: S1. Dry Coptis chinensis until the moisture content is ≤5%, then pulverize it and sieve it to obtain Coptis chinensis powder with a mesh size of 60-80. S2. After heating choline chloride and ethylene glycol in an oil bath and stirring at 80°C for 2.5 hours, the mixture was cooled to 50°C and polyethylene glycol 400 was added. After stirring for another 20 minutes, a eutectic solvent was obtained. The molar ratio of choline chloride, ethylene glycol and polyethylene glycol 400 was 1:1.8:0.8. S3. Mix the eutectic solvent with water and add 1 kg of Coptis chinensis powder. The amount of water added is 5% of the volume of the eutectic solvent. The ratio of Coptis chinensis powder to eutectic solvent is 20 mL / g (that is, 1 g of Coptis chinensis powder is added to 20 mL of eutectic solvent). Extract with pulsed ultrasound at 45℃ for 40 min (ultrasound frequency is 40 kHz, power is 200 W, turn on for 2 seconds and turn off for 1 second). Then, centrifuge at 5000 rpm to obtain the supernatant and filter residue. S4. Repeat step S3 with the filter residue twice, combine the supernatants, and then post-process the supernatant to obtain berberine. The specific post-processing steps are as follows: S4-1. First, adjust the pH of the combined supernatant to 2; S4-2. Then add ammonium sulfate to the supernatant until saturated, and then shake at low temperature for 10 min. S4-3, then add ethyl acetate and n-hexane in a volume ratio of 7:2.5 to prepare a mixed solvent. After standing at low temperature for 30 min, centrifuge to separate the organic phase A, which is mainly composed of berberine, and the aqueous phase B. S4-4. After washing organic phase A with water, the organic solvent was rotary evaporated at 40℃ and -0.09MPa to obtain crude berberine extract. Then, the crude berberine extract was thermally dissolved in hot ethanol at 70℃, filtered, cooled and crystallized, and dried to obtain berberine extract. The eutectic solvent was recovered by vacuum distillation of aqueous phase B at -0.09 MPa and 80 °C.

[0029] Example 3 A method for extracting berberine from Coptis chinensis using a eutectic solvent includes the following steps: S1. Dry Coptis chinensis until the moisture content is ≤5%, then pulverize it and sieve it to obtain Coptis chinensis powder with a mesh size of 60-80. S2. After heating choline chloride and ethylene glycol in an oil bath and stirring at 90°C for 2 hours, the mixture is cooled to 55°C and polyethylene glycol 400 is added. Stirring is continued for 20 minutes to obtain a eutectic solvent. The molar ratio of choline chloride, ethylene glycol and polyethylene glycol 400 is 1:2.2:1.2. S3. Mix the eutectic solvent with water and add 1 kg of Coptis chinensis powder. The amount of water added is 5-8% of the volume of the eutectic solvent. The ratio of Coptis chinensis powder to eutectic solvent is 25 mL / g (that is, 1 g of Coptis chinensis powder is added to 25 mL of eutectic solvent). Extract with pulsed ultrasound at 50℃ for 30 min (ultrasound frequency is 40 kHz, power is 200 W, turn on for 2 seconds and turn off for 1 second). Then centrifuge at 5000 rpm to obtain the supernatant and filter residue. S4. Repeat step S3 with the filter residue twice, combine the supernatants, and then post-process the supernatant to obtain berberine. The specific post-processing steps are as follows: S4-1. First, adjust the pH of the combined supernatant to 3; S4-2. Then add ammonium sulfate to the supernatant until saturated, and then shake at low temperature for 20 minutes. S4-3, then add ethyl acetate and n-hexane in a volume ratio of 7:3.5 to prepare a mixed solvent. After standing at low temperature for 40 min, centrifuge to separate the organic phase A, which is mainly composed of berberine, and the aqueous phase B. S4-4. After washing organic phase A with water, the organic solvent was rotary evaporated at 50℃ and -0.1MPa to obtain crude berberine extract. Then, the crude berberine extract was thermally dissolved in hot ethanol at 70℃, filtered, cooled and crystallized, and dried to obtain berberine extract. The aqueous phase B was distilled under reduced pressure at -0.1 MPa and 100 °C to recover the eutectic solvent.

[0030] Example 4 A method for extracting berberine from Coptis chinensis using a eutectic solvent is provided, following the method in Example 1, except that in step S3, an equal amount of water is replaced with micro / nano bubble water. The micro / nano bubble water is aerated using air via a micro / nano bubble aerator. The micro / nano bubbles in the micro / nano bubble water have a particle size of 200 nm-10 μm and a bubble concentration of 3.6 × 10⁻⁶.6 per mL.

[0031] Example 5 A method for extracting berberine from Coptis chinensis using a eutectic solvent is provided, following the method in Example 1, except that in step S3, an equal amount of water is replaced with micro / nano bubble water. The micro / nano bubble water is aerated using air via a micro / nano bubble aerator. The micro / nano bubbles in the micro / nano bubble water have a particle size of 200 nm-10 μm and a bubble concentration of 1.5 × 10⁻⁶. 7 per mL.

[0032] Example 6 A method for extracting berberine from Coptis chinensis using a eutectic solvent is performed according to the method in Example 1, except that the concentration of berberine in aqueous phase B is >5 mg / mL in step S4-3, and aqueous phase B also undergoes aqueous two-phase extraction and countercurrent partitioning. The specific operations are as follows: Aqueous two-phase extraction: Adjust the pH of aqueous phase B to 2-3, then add a mixed aqueous solution of PEG-6000 and ammonium sulfate, shake and mix for 30 min, then centrifuge to obtain an upper phase containing berberine and PEG and a lower phase containing eutectic solvent and ammonium sulfate. The mass concentration of PEG-6000 in the mixed aqueous solution is 20-25%, and the mass concentration of ammonium sulfate is 15-20%. Countercurrent partitioning: Mix ethyl acetate and n-hexane at a volume ratio of 1:(1-1.1) as the mobile phase, and form a countercurrent extraction with the upper phase obtained from the aqueous two-phase extraction. Specifically: Inject the upper phase into the extraction tube of the countercurrent partitioner, rotate to retain it as the stationary phase, and let the mobile phase flow through the upper phase at a flow rate of 1-5 ml / min. The mobile phase carrying berberine flows out from the end of the column to obtain an organic phase C containing berberine. The remaining liquid is discharged as the aqueous phase D. Step S4-4 is as follows: organic phase C and organic phase D are combined and washed with water, and then rotary evaporated to obtain crude berberine extract. The remaining operations are the same as in Example 1. The aqueous phase D was first evaporated and crystallized to precipitate supersaturated ammonium sulfate crystals. The solution was then filtered, and the filtrate was passed through an ultrafiltration membrane with a molecular weight cutoff of 6000 Da. Finally, the solution was subjected to vacuum distillation to recover the eutectic solvent. The vacuum distillation parameters were the same as in Example 1.

[0033] Example 7 A method for extracting berberine from Coptis chinensis using a eutectic solvent is performed according to the method in Example 6, except that in step S4-3, aqueous phase B undergoes aqueous two-phase extraction and countercurrent partitioning. The specific operations are as follows: Aqueous two-phase extraction: Adjust the pH of aqueous phase B to 2-3, then add a mixed aqueous solution of PEG-6000 and ammonium sulfate, shake and mix for 30 min, then centrifuge to obtain an upper phase containing berberine and PEG and a lower phase containing eutectic solvent and ammonium sulfate. The mass concentration of PEG-6000 in the mixed aqueous solution is 20-25%, and the mass concentration of ammonium sulfate is 15-20%. Countercurrent partitioning: Mix ethyl acetate and n-hexane at a volume ratio of 1:(1-1.1) as the mobile phase, and form a countercurrent extraction with the upper phase obtained from the aqueous two-phase extraction. Specifically: Inject the upper phase into the extraction tube of the countercurrent partitioner, rotate to retain it as the stationary phase, and let the mobile phase flow through the upper phase at a flow rate of 1-5 ml / min. The mobile phase carrying berberine flows out from the end of the column to obtain an organic phase C containing berberine. The remaining liquid is discharged as the aqueous phase D. Step S4-4 is as follows: organic phase C and organic phase D are combined and washed with water, and then rotary evaporated to obtain crude berberine extract. The remaining operations are the same as in Example 1. The aqueous phase D was first evaporated and crystallized to precipitate supersaturated ammonium sulfate crystals. The solution was then filtered, and the filtrate was passed through an ultrafiltration membrane with a molecular weight cutoff of 6000 Da. Finally, the solution was subjected to vacuum distillation to recover the eutectic solvent. The vacuum distillation parameters were the same as in Example 1.

[0034] Example 8 A method for extracting berberine from Coptis chinensis using a eutectic solvent is performed according to the method in Example 6, except that in step S4-3, aqueous phase B undergoes aqueous two-phase extraction and countercurrent partitioning. The specific operations are as follows: Aqueous two-phase extraction: Adjust the pH of aqueous phase B to 2-3, then add a mixed aqueous solution of PEG-6000 and ammonium sulfate, shake and mix for 30 min, then centrifuge to obtain an upper phase containing berberine and PEG and a lower phase containing eutectic solvent and ammonium sulfate. The mass concentration of PEG-6000 in the mixed aqueous solution is 20-25%, and the mass concentration of ammonium sulfate is 15-20%. Countercurrent partitioning: Mix ethyl acetate and n-hexane at a volume ratio of 1:(1-1.1) as the mobile phase, and form a countercurrent extraction with the upper phase obtained from the aqueous two-phase extraction. Specifically: Inject the upper phase into the extraction tube of the countercurrent partitioner, rotate to retain it as the stationary phase, and let the mobile phase flow through the upper phase at a flow rate of 1-5 ml / min. The mobile phase carrying berberine flows out from the end of the column to obtain an organic phase C containing berberine. The remaining liquid is discharged as the aqueous phase D. Step S4-4 is as follows: organic phase C and organic phase D are combined and washed with water, and then rotary evaporated to obtain crude berberine extract. The remaining operations are the same as in Example 1. The aqueous phase D was first evaporated and crystallized to precipitate supersaturated ammonium sulfate crystals. The solution was then filtered, and the filtrate was passed through an ultrafiltration membrane with a molecular weight cutoff of 6000 Da. Finally, the solution was subjected to vacuum distillation to recover the eutectic solvent. The vacuum distillation parameters were the same as in Example 1.

[0035] Comparative Example 1 A method for extracting berberine from Coptis chinensis using a eutectic solvent is performed according to the method in Example 1, except that polyethylene glycol 400 is not added to the eutectic solvent in step S2.

[0036] Comparative Example 2 A method for extracting berberine from Coptis chinensis using a eutectic solvent is carried out according to the method in Example 1, except that polyethylene glycol 400 is not added to the eutectic solvent in step S2, and ethylene glycol is replaced by urea in equal amounts.

[0037] Comparative Example 3 A method for extracting berberine from Coptis chinensis using a eutectic solvent is performed according to the method in Example 1, except that the post-treatment of the supernatant in step S4 is specifically performed as follows: The combined supernatant was first adjusted to pH 2, then cooled to 5°C in an ice-water bath, allowed to stand for 2 hours, cooled to crystallize, filtered, then washed with water and dissolved in 70°C hot ethanol, filtered, cooled to crystallize, and dried to obtain berberine extract.

[0038] Performance testing The purity of the berberine extracts prepared by the methods in the above examples and comparative examples was determined by high performance liquid chromatography (HPLC). The results are shown in Table 1 below, where the extraction rate is ((mass of berberine extract obtained × purity of berberine extract) / (berberine content in coptis powder × mass of coptis powder)) × 100%.

[0039] Table 1: Performance testing Example 1 Example 2 Example 3 Example 4 Example 5 Example 6 Extraction rate / % 86.8 85.4 86.3 89.5 90.1 92.8 purity / % 93.52 92.84 93.17 94.21 94.48 98.8 Continued from Table 1: Performance testing Example 7 Example 8 Comparative Example 1 Comparative Example 2 Comparative Example 3 — Extraction rate / % 91.8 92.5 75.3 72.1 77.6 — purity / % 97.6 97.9 85.6 83.5 80.2 — Referring to the test results in Table 1 above, the eutectic solvent extraction of Coptis chinensis provided in this application has a high extraction rate and high purity for berberine. Combined with the test results of Examples 1, 4, and 5, the extraction rate is higher when micro-nano bubble water is added to the eutectic solvent compared to Example 1. Micro-nano bubble water helps to improve the extraction efficiency. Combined with the test results of Examples 6-8, the aqueous phase after single-stage extraction is further treated by two-phase extraction and countercurrent partitioning to further recover berberine in aqueous phase B, improve the berberine extraction rate, recover the eutectic solvent, and further improve the purity of berberine. Combined with the test results in Comparative Example 1, the extraction rate was significantly reduced when using a traditional binary eutectic solvent for extraction. Combined with the test results in Comparative Example 2, the extraction rate was even lower when using a choline chloride-urea eutectic solvent for extraction. Combined with the test results in Comparative Example 3, the extraction rate was low when the supernatant after extraction with the eutectic solvent was not subjected to the post-processing method in Example 1 of this application, but instead a traditional cooling crystallization method was used, due to problems such as incomplete crystallization and interference from impurities. Using the post-processing method in Example 1 helps to improve the extraction rate.

[0040] This specific embodiment is merely an explanation of this application and is not intended to limit it. After reading this specification, those skilled in the art can make modifications to this embodiment without contributing any inventive step, but such modifications are protected by patent law as long as they fall within the scope of the claims of this application.

Claims

1. A method for extracting berberine from Coptis chinensis using a eutectic solvent, characterized in that, Includes the following steps: S1. After drying, Coptis chinensis is pulverized to obtain Coptis chinensis powder; S2. A eutectic solvent was prepared by mixing choline chloride, ethylene glycol, and polyethylene glycol 400 in a molar ratio of 1:(1.8-2.2):(0.8-1.2). S3. Add Coptis chinensis powder to a eutectic solvent and extract with ultrasonic assistance at 45-50℃ for 30-40 min. After centrifugation, obtain the supernatant and filter residue. S4. Repeat step S3 with the filter residue twice, combine the supernatants, and then post-process the supernatant to obtain berberine.

2. The method for extracting berberine from Coptis chinensis using a eutectic solvent according to claim 1, characterized in that: In step S3, after mixing the eutectic solvent and the micro / nano bubble water, Coptis chinensis powder is added. The amount of micro / nano bubble water added is 5-8% of the volume of the eutectic solvent. The particle size of the micro / nano bubbles in the micro / nano bubble water is 200nm-10μm, and the bubble concentration in the micro / nano bubble water is 10. 6 -10 7 per mL.

3. The method for extracting berberine from Coptis chinensis using a eutectic solvent according to claim 1, characterized in that: In step S3, the ratio of Coptis chinensis powder to eutectic solvent is 20-25 mL / g.

4. The method for extracting berberine from Coptis chinensis using a eutectic solvent according to claim 1, characterized in that: The eutectic solvent in step S2 is prepared by the following method: After stirring choline chloride and ethylene glycol at 80-90℃ for 2-2.5h, the mixture is cooled to 50-55℃, polyethylene glycol 400 is added, and stirring is continued for 20-30min to obtain a eutectic solvent.

5. The method for extracting berberine from Coptis chinensis using a eutectic solvent according to claim 1, characterized in that: The specific procedures for supernatant post-treatment in step S4 are as follows: S4-1. First, adjust the pH of the combined supernatant to 2-3; S4-2. Then add ammonium sulfate to the supernatant until saturated, and then shake at low temperature for 10-20 minutes. S4-3. Add a mixed solvent of ethyl acetate and n-hexane, let stand at low temperature for 30-40 min, and then centrifuge to obtain organic phase A, which is mainly composed of berberine, and aqueous phase B. S4-4. After washing organic phase A with water, the organic solvent is rotary evaporated to obtain crude berberine extract, which is then purified to obtain berberine; the aqueous phase B is distilled under reduced pressure to recover the eutectic solvent.

6. The method for extracting berberine from Coptis chinensis using a eutectic solvent according to claim 5, characterized in that: In step S4-3, the volume ratio of ethyl acetate to n-hexane in the mixed solvent is 7:(2.5-3.5).

7. The method for extracting berberine from Coptis chinensis using a eutectic solvent according to claim 5, characterized in that: The aqueous phase B obtained in step S4-3 is further subjected to aqueous two-phase extraction and countercurrent partitioning, specifically as follows: Aqueous two-phase extraction: The pH of aqueous phase B is adjusted to 2-3, and then a mixed aqueous solution of PEG-6000 and ammonium sulfate is added. The mixture is shaken and then centrifuged to obtain an upper phase containing berberine and PEG and a lower phase containing eutectic solvent and ammonium sulfate. Countercurrent partitioning: Ethyl acetate and n-hexane are mixed in a volume ratio of 1:(1-1.1) as the mobile phase and countercurrently extracted with the upper phase obtained from the aqueous two-phase extraction to obtain an organic phase C containing berberine and an aqueous phase D. Then, step S4-4 is performed, and organic phase C and organic phase A are combined, washed with water and rotary evaporated to obtain crude berberine extract. Aqueous phase D is distilled under reduced pressure to recover the eutectic solvent.

8. The method for extracting berberine from Coptis chinensis using a eutectic solvent according to claim 7, characterized in that: During aqueous two-phase extraction, the mass concentration of PEG-6000 in the mixed aqueous solution is 20-25%, and the mass concentration of ammonium sulfate is 15-20%.

9. A method for extracting berberine from Coptis chinensis using a eutectic solvent according to claim 7, characterized in that: In step S4-4, the aqueous phase D is first evaporated and crystallized to precipitate ammonium sulfate as a supersaturated crystal. After filtration, the filtrate is passed through an ultrafiltration membrane with a molecular weight cutoff of 6000 Da. Finally, the eutectic solvent is recovered by vacuum distillation at -0.09-(-0.1) MPa and 80-100℃.

10. A method for extracting berberine from Coptis chinensis using a eutectic solvent according to claim 7, characterized in that: In step S4-4, the parameters for the rotary evaporation of organic phase C and organic phase A after water washing are: rotary evaporation at 40-50℃ and -0.09-(-0.1) MPa under reduced pressure.