A process for extracting and separating artemisinin

Abstract

The present application relates to the field of plant extraction, and specifically discloses an extraction and separation process of artemisinin. It comprises: taking the raw medicinal material of Artemisia annua and performing pretreatment; using ethylene as a supercritical fluid and acetone as an entrainer, performing supercritical fluid extraction on the pretreated raw medicinal material of Artemisia annua to obtain an Artemisia annua extract, and then mixing the Artemisia annua extract with silica gel after decompression concentration, performing silica gel column separation, and then performing crystallization treatment. The process adopts supercritical fluid extraction technology, using ethylene as a supercritical fluid and acetone as an entrainer to extract and classify the raw medicinal material of Artemisia annua. The extraction performance of ethylene on artemisinin can be accurately controlled by adjusting the temperature and pressure; at the same time, by adding acetone as an entrainer, the solvent characteristics of ethylene are improved, so that it can effectively dissolve and extract low-grade artemisinin under supercritical conditions, and the structure of artemisinin can be avoided from changing, thereby improving the extraction rate and purity of artemisinin.

Description

Technical Field

[0001] The present application relates to the field of drug preparation, and more specifically, to a process for extracting and separating artemisinin. Background Art

[0002] Artemisinin was originally derived from the Chinese herb Artemisia annua, which is used in traditional Chinese medicine to treat symptoms such as malaria and fever. Through systematic research on Artemisia annua, scientists found that its active ingredient artemisinin has a killing effect on malarial parasites. Since then, artemisinin, as a natural product with significant antimalarial activity, has been widely concerned by scientific research and industrial production for its extraction and separation process.

[0003] The core structure of the artemisinin molecule includes a lactone ring and a peroxide bridge, which give artemisinin a powerful antimalarial effect. However, this structure is unstable and is easily destroyed during the separation process, resulting in low extraction efficiency and poor product purity, making it difficult to meet the growing market demand. In addition, traditional artemisinin extraction methods mainly use petroleum ether or n-hexane as extraction solvents, followed by separation and purification by silica gel columns. However, these methods have safety hazards such as low boiling point and flammability and explosion, which are not conducive to scale-up production.

[0004] In recent years, some technologies for extracting artemisinin using supercritical fluid extraction have been developed. Although these extraction processes are relatively environmentally friendly, they basically use carbon dioxide as a supercritical fluid, which is not efficient enough for the extraction and purification of artemisinin. The purity and extraction rate of artemisinin are difficult to meet expectations. In this context, exploring a new process that can maintain the structural stability of artemisinin and significantly improve the extraction efficiency and purity has become an urgent problem to be solved in the industry. Summary of the invention

[0005] In order to improve the extraction efficiency and purity of artemisinin, the present application provides an artemisinin extraction and separation process.

[0006] This application adopts the following technical solutions:

[0007] An artemisinin extraction and separation process comprises the following steps:

[0008] Step S1: taking the raw medicinal material of Artemisia annua and performing pretreatment;

[0009] Step S2: using ethylene as a supercritical fluid and acetone as an entrainer to perform supercritical fluid extraction on the pretreated raw Artemisia annua to obtain an Artemisia annua extract, and then reducing the pressure to concentrate the Artemisia annua extract to remove the fluid and the entrainer to obtain a crude artemisinin extract;

[0010] Step S3: mixing the crude artemisinin extract with silica gel, performing silica gel column separation by dry loading, and collecting the eluate containing artemisinin;

[0011] Step S4: Concentrating the eluate and then subjecting it to crystallization.

[0012] Furthermore, the process parameters of the above-mentioned supercritical fluid extraction are: extraction temperature of 20-23° C., extraction pressure of 11-19 MPa, extraction time of 60-120 min, and extraction agent flow rate of 15-18 kg / h.

[0013] The above technical scheme adopts supercritical fluid extraction technology, uses ethylene as supercritical fluid and acetone as entrainer to extract and classify the raw medicinal material of Artemisia annua. Since the critical temperature and pressure of ethylene are relatively low, the extraction performance of ethylene can be accurately controlled by adjusting the temperature and pressure; at the same time, by adding acetone as an entrainer, the solvent properties of ethylene are improved, so that it can effectively dissolve and extract artemisinin with lower polarity under supercritical conditions, and the structure of artemisinin can be avoided from changing.

[0014] Furthermore, during the supercritical fluid extraction process, the extraction temperature remains unchanged and the extraction pressure is set according to the following procedure:

[0015] The initial pressure is 11-12Mpa. After running smoothly for 10-20 minutes, the first stage of pressurization is carried out to gradually increase the system pressure to 15-16Mpa, and it runs at this pressure for 40-60 minutes. Then, the second stage of pressurization is carried out to gradually increase the system pressure to 18-19Mpa, and it runs at this pressure for 40-70 minutes.

[0016] Furthermore, the pressure increasing rate of the first stage of pressure increasing is 0.4-0.6Mpa / min; the pressure increasing rate of the second stage of pressure increasing is 0.2-0.3Mpa / min.

[0017] In the above technical scheme, by adopting the extraction process of heat preservation and variable pressure, the polarity and solubility of the extractant are adjusted by changing the pressure. During the extraction process, substances of different polarities are extracted in order at different stages, which not only improves the extraction efficiency of artemisinin, but also reduces the content of other non-artemisinin substances in the crude extract of artemisinin, and can be effectively separated during the extraction process, thereby making the subsequent purification step easier.

[0018] Furthermore, in the above-mentioned silica gel column separation process of the crude artemisinin extract, a gradient elution method is adopted, firstly washing with a n-hexane-petroleum ether mixed system for 2-3 column volumes; and then washing with an ethyl acetate-petroleum ether mixed system for 4-5 column volumes.

[0019] Furthermore, the content of n-hexane in the n-hexane-petroleum ether mixed system is 15-18%, and the content of ethyl acetate in the ethyl acetate-petroleum ether mixed system is 9-14%.

[0020] The crude artemisinin extract is purified and separated by silica gel column chromatography. Dry loading is performed and gradient elution is performed using a n-hexane-petroleum ether mixed system and an ethyl acetate-petroleum ether mixed system as eluents. This can effectively separate the artemisinin in the crude artemisinin extract, improve the purity of the artemisinin, and facilitate crystallization to obtain a product with a purity greater than 99%.

[0021] Furthermore, before the above-mentioned silica gel column separation process of the crude artemisinin extract, the following color removal and impurity removal steps are also included: the crude artemisinin extract is dissolved in an ethyl acetate-petroleum ether mixture system to obtain a crude extract; the crude extract is evenly mixed with a 0.5-0.8% polyacrylamide solution, allowed to stand for 6-9 hours, centrifuged and washed, and then heated in a water bath at 50-60°C for 1-2 hours for desorption.

[0022] In this technical solution, before the crude artemisinin extract is subjected to silica gel column separation, the crude artemisinin extract is mixed with polyacrylamide, which can form a water-soluble polymer with strong adsorption and precipitation properties, and can adsorb pigments and highly polar impurities in the crude extract. After adsorption, the adsorption force between the target molecule and the polyacrylamide can be reduced by heating to perform desorption.

[0023] Furthermore, the above method for pre-treating the raw medicinal material of Artemisia annua includes: drying the raw medicinal material of Artemisia annua at 55-60° C., crushing it through a 20-mesh sieve, and homogenizing the obtained powder.

[0024] Drying, crushing and homogenizing the raw Artemisia annua materials can help to make the contact between the raw Artemisia annua materials and the extractant more uniform, thereby improving the extraction efficiency.

[0025] In summary, this application has the following beneficial effects:

[0026] The artemisinin extraction and classification process provided in the present application adopts supercritical fluid extraction technology, uses ethylene as a supercritical fluid, and uses acetone as an entrainer to extract and classify the original Artemisia annua medicinal material. Since the critical temperature and pressure of ethylene are relatively low, the extraction performance of ethylene on artemisinin can be accurately controlled by adjusting the temperature and pressure; at the same time, by adding acetone as an entrainer, the solvent properties of ethylene are improved, so that it can effectively dissolve and extract artemisinin with lower polarity under supercritical conditions, and the structure of artemisinin can be avoided from changing, thereby improving the extraction rate and purity of artemisinin. DETAILED DESCRIPTION

[0027] The embodiments of the present invention will be described in detail below in conjunction with examples. However, those skilled in the art will understand that the following examples are only used to illustrate the present invention and should not be construed as limiting the scope of the present invention. The specific conditions not specified in the examples are carried out according to conventional conditions or conditions recommended by the manufacturer. The reagents or instruments used without indicating the manufacturer are all conventional products that can be purchased commercially.

[0028] The specific embodiments of the present invention are described in detail below. It should be understood that the specific embodiments described herein are only used to illustrate and explain the present invention, and are not used to limit the present invention.

[0029] Example 1

[0030] This embodiment provides a process for extracting and separating artemisinin, which comprises the following steps:

[0031] 1. Dry the raw Artemisia annua at 55-60°C, grind it through a 20-mesh sieve, and homogenize the resulting powder to obtain Artemisia annua powder;

[0032] 2. Put the Artemisia annua powder into an extraction kettle, use ethylene as a supercritical fluid and acetone as an entrainer (content of 6%) to perform supercritical fluid extraction: the extraction temperature is 22°C, the extraction pressure is 15Mpa, the extraction time is 90min, and the flow rate of the extractant is 17kg / h to obtain an Artemisia annua extract. The Artemisia annua extract is concentrated under reduced pressure to remove the supercritical fluid and the entrainer to obtain a crude artemisinin extract.

[0033] 3. The crude artemisinin extract was mixed with 2.5 times the weight of silica gel, and the mixture was dry-loaded for silica gel column separation and gradient elution. The elution method was as follows: 2-3 column volumes were washed with a mixed system of n-hexane-petroleum ether (the content of n-hexane was 15-18%), and 4-5 column volumes were washed with a mixed system of ethyl acetate-petroleum ether (the content of ethyl acetate was 9-14%). The fractions in the middle were collected. After TLC analysis, the fractions containing artemisinin were mixed and concentrated under reduced pressure to obtain artemisinin extract.

[0034] 4. The obtained artemisinin extract was resuspended in tetrahydrofuran at a mass-to-volume ratio of 1:8 for pure crystallization.

[0035] Example 2

[0036] This embodiment provides an extraction and separation process of artemisinin, which is different from that of embodiment 1 in that the process parameters of supercritical fluid extraction are different:

[0037] The extraction temperature was 20° C., the extraction pressure was 19 MPa, the extraction time was 60 min, the extraction agent flow rate was 18 kg / h, and the entrainer content was 5%.

[0038] Example 3

[0039] This embodiment provides an extraction and separation process of artemisinin, which is different from that of embodiment 1 in that the process parameters of supercritical fluid extraction are different:

[0040] The extraction temperature was 23° C., the extraction pressure was 11 MPa, the extraction time was 120 min, the extraction agent flow rate was 15 kg / h, and the entrainer content was 8%.

[0041] Example 4

[0042] This embodiment provides an extraction and separation process of artemisinin, which is different from Embodiment 1 in that the supercritical fluid extraction process adopts a heat preservation and pressure increase process, specifically:

[0043] The temperature of supercritical fluid extraction was 22°C, the initial pressure was 11 MPa, and it ran smoothly for 15 minutes; the first stage of pressurization was carried out at a pressure increase rate of 0.5 MPa / min, and the system pressure was gradually increased to 16 MPa, and it ran at this pressure for 45 minutes; then the second stage of pressurization was carried out at a pressure increase rate of 0.2 MPa / min, and the system pressure was gradually increased to 19 MPa, and it ran at this pressure for 60 minutes to obtain the Artemisia annua extract.

[0044] Example 5

[0045] This embodiment provides an extraction and separation process of artemisinin, which is different from Embodiment 1 in that the supercritical fluid extraction process adopts a heat preservation and pressure increase process, specifically:

[0046] The temperature of supercritical fluid extraction is 22°C, the initial pressure is 12 MPa, and it runs smoothly for 20 minutes; the first stage of pressurization is carried out at a pressure increase rate of 0.4 MPa / min, and the system pressure is gradually increased to 15 MPa, and it runs at this pressure for 60 minutes; then the second stage of pressurization is carried out at a pressure increase rate of 0.3 MPa / min, and the system pressure is gradually increased to 19 MPa, and it runs at this pressure for 40 minutes to obtain the Artemisia annua extract.

[0047] Example 6

[0048] This embodiment provides an extraction and separation process of artemisinin, which is different from Embodiment 1 in that the supercritical fluid extraction process adopts a heat preservation and pressure increase process, specifically:

[0049] The temperature of supercritical fluid extraction was 22°C, the initial pressure was 11 MPa, and it ran smoothly for 10 minutes; the first stage of pressurization was carried out at a pressure increase rate of 0.6 MPa / min, and the system pressure was gradually increased to 16 MPa, and it ran at this pressure for 40 minutes; then the second stage of pressurization was carried out at a pressure increase rate of 0.2 MPa / min, and the system pressure was gradually increased to 18 MPa, and it ran at this pressure for 65 minutes to obtain the Artemisia annua extract.

[0050] Comparative Examples 1-5

[0051] The difference between this comparative example and Example 1 is that the solvent and process parameters of supercritical fluid extraction are different:

[0052] Comparative Example 1: Using supercritical ethylene fluid as the extractant, wherein the supercritical ethylene fluid does not contain an entrainer, that is, does not contain acetone;

[0053] Comparative Example 2: Supercritical ethylene fluid is used as the extractant, and the extractant contains 6% isopropanol as an entrainer.

[0054] Comparative Example 3: Using supercritical CO2 as the extraction agent, the process parameters are: extraction time 6h, extraction temperature 50°C, extraction pressure 45Mpa, and flow rate 25kg / h.

[0055] Comparative Example 4: Using supercritical ethylene fluid as the extractant, the extractant contains 6% acetone as the entrainer, the extraction pressure is 9 MPa, and other conditions are the same;

[0056] Comparative Example 5: Supercritical ethylene fluid is used as the extractant, the extractant contains 6% acetone as the entrainer, the extraction pressure is 21 MPa, and other conditions are the same.

[0057] The extraction and separation processes provided in Examples 1 and 4 and Comparative Examples 1-5 were investigated based on the yield of the crude artemisinin extract after supercritical fluid extraction. The results are shown in Table 1:

[0058] Table 1.

[0059]

[0060] As can be seen from Table 1, the extraction and separation process of artemisinin provided in Example 1 of the present application has a yield of a crude artemisinin extract in the supercritical extraction step that is better than that of Comparative Examples 1-5, especially better than Comparative Example 3. In Comparative Example 3, commonly used supercritical CO2 is used as an extractant, and the optimal parameter combination obtained by orthogonal design is used. The yield of the crude artemisinin extract obtained is 4.39%. Although it is not much different from Example 1 of the present application, it should be emphasized that the extraction time of Example 3 is 6h, while that of Example 1 is only 1.5h, which can greatly improve the extraction efficiency and reduce the equipment operation cost. In addition, compared with Comparative Examples 1 and 2, the addition of entrainer acetone in the supercritical fluid can further improve the yield of the crude artemisinin extract.

[0061] Compared with Example 1, Example 4 adopts the method of heat preservation and pressure increase for extraction, and the total yield of the crude artemisinin extract obtained is as high as 5.27%, which shows that this heat preservation and pressure increase process is more conducive to the efficient dissolution and extraction of artemisinin.

[0062] Example 7

[0063] This embodiment provides an extraction and separation process of artemisinin, which is different from embodiment 4 in that: before the artemisinin crude extract is subjected to silica gel column separation, the following color removal and impurity removal steps are also included:

[0064] The crude artemisinin extract was dissolved in 100 ml of an ethyl acetate-petroleum ether mixture (the content of ethyl acetate was 8%) to obtain a crude extract with a mass fraction of 12 wt%; the crude extract was mixed evenly with a 0.6% polyacrylamide solution, allowed to stand for 7 h, centrifuged and washed, and then heated in a water bath at 55° C. for 1.5 h for desorption. After recovering the solvent, the obtained concentrate was subjected to silica gel column chromatography.

[0065] Example 8

[0066] This embodiment provides an extraction and separation process of artemisinin, which is different from embodiment 4 in that: before the artemisinin crude extract is subjected to silica gel column separation, the following color removal and impurity removal steps are also included:

[0067] The crude artemisinin extract was dissolved in 100 ml of an ethyl acetate-petroleum ether mixture (the content of ethyl acetate was 8%) to obtain a crude extract with a mass fraction of 10 wt %; the crude extract was mixed evenly with a 0.5% polyacrylamide solution, allowed to stand for 9 h, centrifuged and washed, and then heated in a water bath at 60° C. for 1 h for desorption. After recovering the solvent, the obtained concentrate was subjected to silica gel column chromatography.

[0068] Example 9

[0069] This embodiment provides an extraction and separation process of artemisinin, which is different from embodiment 4 in that: before the artemisinin crude extract is subjected to silica gel column separation, the following color removal and impurity removal steps are also included:

[0070] The crude artemisinin extract was dissolved in 100 ml of an ethyl acetate-petroleum ether mixture (the content of ethyl acetate was 8%) to obtain a crude extract with a mass fraction of 14 wt %; the crude extract was mixed evenly with a 0.8% polyacrylamide solution, allowed to stand for 6 h, centrifuged and washed, and then heated in a water bath at 50° C. for 2 h for desorption. After recovering the solvent, the obtained concentrate was subjected to silica gel column chromatography.

[0071] Comparative Example 6

[0072] The difference between this comparative example and Example 4 is that activated carbon is used as an adsorbent for removing color and impurities.

[0073] Comparative Examples 7-10

[0074] The difference between this comparative example and Example 4 is that the conditions for removing color and impurities using polyacrylamide are different:

[0075] Comparative Example 7: The concentration of polyacrylamide is 0.5%;

[0076] Comparative Example 8: The concentration of polyacrylamide is 1.0%;

[0077] Comparative Example 9: The desorption temperature was 45°C and the time was 1.5 h;

[0078] Comparative Example 10: The desorption temperature was 65°C and the desorption time was 1.5 h.

[0079] The extraction and separation processes provided in Examples 1, 4, 7 and Comparative Examples 6-10 were investigated based on the yield and purity of artemisinin obtained after pure crystallization. The results are shown in Table 2:

[0080] Table 2.

[0081]

[0082]

[0083] As can be seen from Table 2, when the color removal and impurity removal step is added before silica gel column chromatography, the product purity can be significantly improved without significantly reducing the yield of artemisinin. At the same time, compared with activated carbon and polyacrylamide, although the product purity of Comparative Example 6 has increased to a certain extent after the color removal and impurity removal with activated carbon, a part of artemisinin is lost, and the yield is reduced. Similarly, when polyacrylamide is used as an adsorbent, the content of the adsorbent and the temperature of desorption will affect the product yield. This specific embodiment is only an explanation of the present application, and it is not a limitation of the present application. After reading this specification, those skilled in the art can make modifications to the present embodiment without creative contribution as needed, but as long as they are within the scope of the claims of the present application, they are protected by the patent law.

Claims

1. A process for extracting and separating artemisinin, characterized in that: It includes: Take the raw medicinal material of Artemisia annua and perform pretreatment; Using ethylene as a supercritical fluid and acetone as an entrainer, the pretreated raw Artemisia annua extract is subjected to supercritical fluid extraction to obtain an Artemisia annua extract, and the Artemisia annua extract is concentrated under reduced pressure to remove the supercritical fluid and the entrainer to obtain a crude artemisinin extract; The process parameters of the supercritical fluid extraction are as follows: extraction temperature is 20-23°C, extraction time is 60-120min, extraction agent flow rate is 15-18kg / h, and entrainer addition amount is 5-8%; during the supercritical fluid extraction process, the extraction temperature remains unchanged, and the extraction pressure is set according to the following procedure: the initial pressure is 11-12Mpa, and after running steadily for 10-20min; the first stage of pressurization is performed, the system pressure is gradually increased to 15-16Mpa, and the system is operated at this pressure for 40-60min; then the second stage of pressurization is performed, the system pressure is gradually increased to 18-19Mpa, and the system is operated at this pressure for 40-70min; The artemisinin crude extract is dissolved in an ethyl acetate-petroleum ether mixture to obtain a crude extract; the crude extract is mixed evenly with a 0.5-0.8% polyacrylamide solution, allowed to stand for 6-9 hours, centrifuged and washed, and then heated in a water bath at 50-60° C. for 1-2 hours for desorption; The desorbed product is mixed with silica gel, and separated on a silica gel column by dry loading, and the eluate containing artemisinin is collected; The eluate is concentrated and then subjected to crystallization.

2. The process for extracting and separating artemisinin according to claim 1, characterized in that: The pressure increasing rate of the first stage of supercharging is 0.4-0.6Mpa / min; the pressure increasing rate of the second stage of supercharging is 0.2-0.3Mpa / min.

3. The process for extracting and separating artemisinin according to claim 1, characterized in that: During the silica gel column separation of the crude artemisinin extract, a gradient elution method is adopted, firstly washing with a n-hexane-petroleum ether mixture system for 2-3 column volumes; and then washing with an ethyl acetate-petroleum ether mixture system for 4-5 column volumes.

4. The process for extracting and separating artemisinin according to claim 3, characterized in that: The content of n-hexane in the n-hexane-petroleum ether mixed system is 15-18%, and the content of ethyl acetate in the ethyl acetate-petroleum ether mixed system is 9-14%.

5. The process for extracting and separating artemisinin according to claim 1, characterized in that: The method for pretreating the raw Artemisia annua medicinal material comprises: drying the raw Artemisia annua medicinal material at 55-60° C., crushing the raw Artemisia annua medicinal material and passing it through a 20-mesh sieve, and homogenizing the obtained powder.

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