WO2020220852A1 - 从大麻中提取大麻二酚的方法 - Google Patents

从大麻中提取大麻二酚的方法 Download PDF

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WO2020220852A1
WO2020220852A1 PCT/CN2020/079693 CN2020079693W WO2020220852A1 WO 2020220852 A1 WO2020220852 A1 WO 2020220852A1 CN 2020079693 W CN2020079693 W CN 2020079693W WO 2020220852 A1 WO2020220852 A1 WO 2020220852A1
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hemp
extraction
water
ethanol
present
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PCT/CN2020/079693
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French (fr)
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于朝晖
常坦然
高伟博
柳旭
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云南汉盟制药有限公司
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C37/00—Preparation of compounds having hydroxy or O-metal groups bound to a carbon atom of a six-membered aromatic ring
    • C07C37/004—Preparation of compounds having hydroxy or O-metal groups bound to a carbon atom of a six-membered aromatic ring by obtaining phenols from plant material or from animal material
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C37/00—Preparation of compounds having hydroxy or O-metal groups bound to a carbon atom of a six-membered aromatic ring
    • C07C37/50—Preparation of compounds having hydroxy or O-metal groups bound to a carbon atom of a six-membered aromatic ring by reactions decreasing the number of carbon atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C37/00—Preparation of compounds having hydroxy or O-metal groups bound to a carbon atom of a six-membered aromatic ring
    • C07C37/68—Purification; separation; Use of additives, e.g. for stabilisation
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C37/00—Preparation of compounds having hydroxy or O-metal groups bound to a carbon atom of a six-membered aromatic ring
    • C07C37/68—Purification; separation; Use of additives, e.g. for stabilisation
    • C07C37/685—Processes comprising at least two steps in series
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C37/00—Preparation of compounds having hydroxy or O-metal groups bound to a carbon atom of a six-membered aromatic ring
    • C07C37/68—Purification; separation; Use of additives, e.g. for stabilisation
    • C07C37/70—Purification; separation; Use of additives, e.g. for stabilisation by physical treatment
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C37/00—Preparation of compounds having hydroxy or O-metal groups bound to a carbon atom of a six-membered aromatic ring
    • C07C37/68—Purification; separation; Use of additives, e.g. for stabilisation
    • C07C37/70—Purification; separation; Use of additives, e.g. for stabilisation by physical treatment
    • C07C37/82—Purification; separation; Use of additives, e.g. for stabilisation by physical treatment by solid-liquid treatment; by chemisorption
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C37/00—Preparation of compounds having hydroxy or O-metal groups bound to a carbon atom of a six-membered aromatic ring
    • C07C37/68—Purification; separation; Use of additives, e.g. for stabilisation
    • C07C37/70—Purification; separation; Use of additives, e.g. for stabilisation by physical treatment
    • C07C37/84—Purification; separation; Use of additives, e.g. for stabilisation by physical treatment by crystallisation
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C39/00—Compounds having at least one hydroxy or O-metal group bound to a carbon atom of a six-membered aromatic ring
    • C07C39/23—Compounds having at least one hydroxy or O-metal group bound to a carbon atom of a six-membered aromatic ring polycyclic, containing six-membered aromatic rings and other rings, with unsaturation outside the aromatic rings
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C2601/00—Systems containing only non-condensed rings
    • C07C2601/12—Systems containing only non-condensed rings with a six-membered ring
    • C07C2601/16—Systems containing only non-condensed rings with a six-membered ring the ring being unsaturated

Definitions

  • the invention relates to the field of medical technology, in particular to a method for extracting cannabidiol from cannabis.
  • Industrial hemp (scientific name: Cannabis sativa L.) is a plant of the moraceae and cannabis genus.
  • the tetrahydrocannabinol (THC) content in the mosaic during the growth period is less than three thousandths, and it does not have the value of extracting tetrahydrocannabinol or its direct use Drug use can be legally grown on a large scale and industrialized, and its economic and medicinal value is extremely high.
  • Cannabidiol compounds are a class of active substances contained in cannabis plants, mainly including tetrahydrocannabinol (THC), tetrahydrocannabinol (THCV), cannabidiol (CBD), cannabidiol (CBG), and cannabis Diphenols (CBDV), etc., these five account for more than 90% of cannabidiol compounds.
  • THC tetrahydrocannabinol
  • THCV tetrahydrocannabinol
  • CBD cannabidiol
  • CBG cannabidiol
  • CBDV cannabis Diphenols
  • Cannabidiol (CBD, cannabidiol), one of the most important non-addictive components in plants, has antispasmodic, anti-rheumatoid arthritis, anti-anxiety and other pharmacological activities, and can hinder tetrahydrocannabinol (THC) against The adverse effects of the human nervous system have become a hot spot for drug development.
  • CBD cannabidiol
  • Patent PCT/CN2017/071993 mentions a method of extracting cannabidiol from cannabis: the extraction part of cannabis is crushed, dried and processed to obtain medicinal powder, which is extracted with 30-100% (V/V) ethanol. Obtain the extract, concentrate under reduced pressure to obtain the extract, add water while hot for precipitation to remove impurities in the water layer, add 10-100% (V/V) ethanol to the precipitate to dissolve, and apply to column chromatography to obtain CBD-rich The target product is then crystallized with organic solvents such as ethanol, washed and dried to obtain a finished product of cannabidiol with a purity of about 99%.
  • organic solvents such as ethanol
  • the crystallization is carried out at low temperature or normal temperature and pressure, standing for 12-72h. Due to the amplification effect, the purity of different crystal parts (such as crystal surface, tank side wall and bottom) of a single crystallization is different Larger, after mixing, the content has not reached more than 99%, and recrystallization is required; in addition, the crystals formed in this way are usually large and very compact, which is difficult to harvest and crush, which is no longer suitable for the requirements of large-scale production.
  • the present invention provides a method for extracting cannabidiol from hemp.
  • the invention provides a method for preparing injection-grade cannabidiol raw materials to solve the problems of high loss and low efficiency of the original process, and fundamentally solve the problem of crystallization industrialization, and at the same time, make the CBD finished product meet the requirements of injection-grade raw materials.
  • the present invention provides a method for extracting cannabidiol from cannabis, including the following steps:
  • Step 1 Take hemp, crush and dry, to obtain hemp powder
  • Step 2 Take the cannabis powder, collect the extract after alcohol extraction, first concentrate, mix with water, decarboxylate, and then concentrate the second to obtain an extract; the temperature of the decarboxylation is 60°C ⁇ 130°C, and the time is 1h ⁇ 3h ;
  • Step 3 Take the extract and mix it with water, adjust the pH to 3.5-6.5, sink with water, centrifuge, collect the precipitate, mix with ethanol, purify, and obtain a concentrated solution through the third concentration; then decolorize, desensitize, and filter , The fourth concentration, to obtain a thick paste;
  • Step 4 Take the thick paste and mix it with solvent at 10°C to 80°C, continue stirring at -10°C to 20°C to crystallize for 12h to 72h, wash and dry.
  • the hemp is one or a combination of two or more of industrial hemp, intermediate hemp or medicinal hemp.
  • the extraction part of the hemp is one or a combination of two or more of hemp flower, hemp leaf, hemp root, hemp stalk core, or hemp seed meal.
  • the step 1 is pulverized to 10-80 mesh.
  • the alcohol extraction in step 2 adopts 30-100% (V/V) ethanol extraction, and the extraction includes reflux extraction, ultrasonic extraction and/or soak extraction; the alcohol extraction The number of times is 1 to 3 times.
  • the reflux extraction includes: using 2-10 times the amount of the medicinal material to perform reflux extraction for 1 to 3 times, each for 0.5 to 3 hours;
  • the ultrasonic extraction includes: using 2 to 10 times the amount of the medicinal material to perform ultrasonic extraction for 1 to 3 times, each time for 0.1 to 1 h;
  • the soaking extraction includes: using 2-10 times the amount of the medicinal material to soak and extract 1 to 3 times, each for 0.5-5h.
  • the first concentration or the second concentration in step 2 is concentrated under reduced pressure at 50-70°C to a relative density of 1.05-1.35 (measured at 50°C).
  • the amount of water added in step 2 is 1/10 to 1/100 times the amount (W/V) of the medicinal material.
  • the pH adjustment adopts organic acid, inorganic acid or base, which may include one or more of formic acid, acetic acid, oxalic acid, hydrochloric acid, nitric acid, sulfuric acid and the like. More preferred are formic acid, oxalic acid and hydrochloric acid.
  • the alkali is sodium hydroxide, potassium hydroxide or ammonia water, more preferably sodium hydroxide.
  • the amount of water in step 3 is 1-10 times the amount of medicinal materials, the temperature of the water settling is 0°C to 20°C, and the time of the water settling is 1 to 48h.
  • the rotation speed of the centrifugation is 4000 rpm to 10000 rpm, and the time of the centrifugation is 3 minutes to 30 minutes.
  • the mass-volume ratio of the precipitate to ethanol in step 3 is 4: (1 to 4); the purification is column chromatography of a macroporous ion exchange resin, and the column layer
  • the elution solvent is used for gradient elution; the gradient elution is: removing impurities with 0.05% to 2% (M/M) alkaline water, and then using 60% to 80% (V/V) of pH 2 to 5.
  • the alkaline water for gradient elution is an aqueous solution of sodium hydroxide, potassium hydroxide or ammonia, more preferably an aqueous solution of sodium hydroxide
  • the filler of the chromatography column includes strong base No. 4, strong base DKx4, Shanghai 763 # , # 702 weak base resin, D301, D303, D380, D301K, macroporous weak base resin One or more.
  • the step of gradient elution includes first removing impurities with 0.05% to 2% (M/M) alkaline water, and then using 60% to 80% of pH 2 to 5 (V /V) acid ethanol is eluted to obtain the target product part, and finally the column is flushed with 90-95% (V/V) acid ethanol of pH 2 to 5 to regenerate the chromatography column.
  • the above-mentioned gradient elution step not only makes the target product partly high purity and good color, but also makes the chromatographic column continuously regenerate and can be recycled.
  • activated carbon is used for the decolorization and desensitization in step 3, and the added amount of activated carbon is 0.2% to 0.5% (W/W) of the mass of cannabidiol in the concentrate, so
  • the temperature for decolorization and desensitization is 45° C. to 75° C., and the time for decolorization and desensitization is 0.5 h to 1 h;
  • the filtration is pressure filtration or suction filtration, and the filter mesh used for the filtration has a pore size of not less than 400 mesh.
  • the activated carbon is in powder form or granules of 0.05 mm to 0.5 mm.
  • the solvent in step 4 is one or a combination of two or more of butane, pentane, hexane, heptane, ethyl acetate, acetone or ethanol; the washing Wash with water or 5%-40% (V/V) ethanol at 0°C ⁇ 24°C; the drying includes one or more of spray drying, vacuum drying, freeze drying, near infrared drying or microwave drying, so The drying temperature is 30°C to 65°C.
  • the drying temperature is 40°C to 55°C.
  • a pulverization step is further included.
  • the pulverization includes steam flow pulverization and/or freeze pulverization, and the pulverization temperature is 0°C to 65°C.
  • the solvent is one or a combination of two or more of hexane, heptane, ethyl acetate, acetone, or ethanol.
  • Ethanol (or acetone) here refers to an ethanol-water (or acetone-water) system, and the crystal concentration is 5%-65% (V/V), more preferably 10%-25% (V/V).
  • the present invention extracts cannabidiol (CBD) from cannabis, and during continuous amplification, the process route is fully optimized: 1) After extraction, the conversion decarboxylation can make the decarboxylation more complete (see Figure 6); 2) the acid precipitation greatly improves the cannabis II The yield of phenol (CBD); 3) Using macroporous ion exchange resin column chromatography, the impurity removal effect is better.
  • CBD cannabidiol
  • the dynamic crystallization process with scraped blades and specific solvents has higher crystallization efficiency and is suitable for large-scale Production (see Figure 7), and the physical properties of the obtained crystals are relatively better, which is conducive to subsequent formulation development; 5)
  • the CBD raw materials prepared by the method of the present invention can meet the requirements of injection level, extending the scope of its modern application.
  • Figure 1 shows the chromatogram of CBD standard (sigma).
  • FIG. 2 shows the chromatogram of CBD product 1
  • Figure 3 shows a chromatogram of CBD product 2
  • Figure 4 shows a chromatogram of CBD product 3
  • Figure 5 shows a chromatogram of CBD product 4.
  • Figure 6 shows the chromatogram of the loading solution; among them, Figure 6(A) shows the product 4 prepared in the comparative example (first decarboxylation followed by extraction), and Figure 6(B) shows the product 1 prepared in Example 1 of the present invention. Extract first and then decarboxylate);
  • Fig. 7 shows a crystal sample diagram; among them, Fig. 7(A) to Fig. 7(B) show a crystal sample of Example 1 of the present invention; Fig. 7(C) to Fig. 7(E) are a crystal sample of a comparative example.
  • the invention discloses a method for extracting cannabidiol from hemp, and those skilled in the art can learn from the content of this article and appropriately improve the process parameters to achieve it.
  • all similar substitutions and modifications are obvious to those skilled in the art, and they are all deemed to be included in the present invention.
  • the method and application of the present invention have been described through the preferred embodiments. It is obvious that relevant personnel can modify or appropriately change and combine the methods and applications described herein without departing from the content, spirit and scope of the present invention to achieve and Apply the technology of the present invention.
  • the invention aims to provide a method for preparing injection-grade cannabidiol raw materials to solve the problems of high loss and low efficiency of the original process, and at the same time, to push the quality of CBD finished products to the height of injection-grade.
  • the present invention provides a method for extracting cannabidiol from cannabis, the method comprising the following steps:
  • the extract is concentrated under reduced pressure to no alcohol, ethanol is recovered, 1/10 to 1/100 times (W/V) of purified water is added to the medicinal material, decarboxylated, and concentrated under normal pressure for 1 to 2 hours to obtain an extract; the decarboxylation
  • the temperature is 60°C ⁇ 130°C, and the time is 1h ⁇ 3h;
  • step 4) Concentrate the decolorization and desensitization solution of step 7) to an anhydrous thick paste, add a specific solvent to supersaturate and dissolve, put the supernatant in a crystallization tank, cool to room temperature, continue to stir and crystallize for 12 to 72 hours, and finally obtain the particle size Homogeneous crystals
  • the crystals are washed with purified water or ethanol to obtain the initial product;
  • the obtained preliminary product is dried and crushed by a suitable drying method to obtain the finished product of cannabidiol.
  • the hemp of the present invention is selected from one or a combination of two or more of industrial hemp, intermediate hemp or medicinal hemp.
  • the extraction part in step 1) of the present invention is selected from one or a combination of two or more of hemp flower, hemp leaf, hemp root, hemp stalk core and hemp seed meal.
  • the extracted part in step 1) of the present invention is hemp flower and hemp leaf.
  • the cannabis extraction part in step 1) of the present invention is crushed to 10-80 mesh, for example, 10 mesh, 12 mesh, 14 mesh, 16 mesh, 18 mesh, 20 mesh, 25 mesh, 30 mesh , 35 mesh, 40 mesh, 45 mesh, 50 mesh, 60 mesh, 70 mesh, 80 mesh, etc.; further preferably 20-60 mesh; more preferably 25-50 mesh; most preferably 40 mesh.
  • the cannabis flowers and leaves are crushed to the aforementioned mesh number range, so that cannabidiol can be fully extracted in the subsequent ethanol extraction step.
  • the amount of ethanol in step 2) of the present invention is 4-10 times the amount of medicinal materials, for example, it can be 4 times, 4.5 times, 5 times, 6 times, 7 times, 8 times, 9 times, 10 times. Times the amount of medicinal materials; further preferably 4-8 times the amount of medicinal materials; more preferably 4-6 times the amount of medicinal materials.
  • the number of times of ethanol extraction in step 2) of the present invention is 1 to 3 times.
  • the extraction method of ethanol extraction in step 2) of the present invention is reflux extraction, ultrasonic extraction and/or soak extraction.
  • the time for the reflux extraction is 0.5-3h per extraction; the time for the ultrasonic extraction is 0.1-1h per extraction; and the time for soaking extraction is 0.5-5h per extraction.
  • the step 2) is: extracting the medicinal powder with 4-8 times the amount of the medicinal material with 30-100% (V/V) ethanol for 1 to 3 times to obtain the extract.
  • the relative density measured when the extract is concentrated to 50°C in step 2) of the present invention is 1.05 to 1.35.
  • the amount of water used for precipitation in step 3) of the present invention is 1 to 10 times the amount of medicinal materials, for example, it can be 1 time, 1.1 times, 1.2 times, 1.3 times, 1.4 times, 1.5 times, 1.6 times , 1.7 times, 1.8 times, 1.9 times, 2 times, 2.25 times, 2.5 times, 3 times, 3.5 times, 4 times, 4.5 times, 5 times, 5.5 times, 6 times, 6.5 times, 7 times, 7.5 times, 8 Times, 8.5 times, 9 times, 9.5 times, and 10 times the amount of medicinal materials; more preferably 2-8 times the amount of medicinal materials; more preferably 3-6 times the amount of medicinal materials.
  • the acid for adjusting pH by precipitation in step 3) of the present invention is one or more of formic acid, acetic acid, oxalic acid, hydrochloric acid, nitric acid, sulfuric acid and the like. More preferred are formic acid, oxalic acid and hydrochloric acid.
  • the alkali is sodium hydroxide, potassium hydroxide or ammonia water, more preferably sodium hydroxide.
  • the pH of the sedimentation adjustment is 2.5-6.5, for example 2.5, 2.7, 2.9, 3.0, 3.3, 3.5, 3.7, 3.9, 4.0, 4.5, 5.0, 5.5, 6.0, 6.5, etc., more preferably 3.5 to 5.5.
  • the temperature of the water settling in step 3) of the present invention is 0-20°C, for example, it can be 0°C, 0.1°C, 0.5°C, 0.8°C, 1.0°C, 1.5°C, 1.75°C, 2°C , 3°C, 4°C, 5°C, 6°C, 7°C, 8°C, 9°C, 10°C, 11°C, 13°C, 15°C, 17°C, 18°C, 19°C, 20°C, etc.; more preferably 2 to 18°C; more preferably 5 to 15°C.
  • the water settling time in step 3) of the present invention is 1-48h, for example, it can be 1h, 1.1h, 1.3h, 1.5h, 1.9h, 2h, 2.5h, 3h, 4h, 5h, 6h, 7h, 8h, 9h, 10h, 15h, 20h, 30h, 40h, 48h, etc.; more preferably 2-36h; more preferably 5-30h.
  • the step 3) is: the extract is submerged with 1-10 times the amount of purified water of the medicinal material at a temperature of 0-20°C for 1 to 48 hours to remove impurities to obtain Water sinking liquid.
  • the filler of the chromatography column used in the column chromatography in step 3) of the present invention includes but not limited to strong base No. 4, strong base DKx4, Shanghai 763#, #702 weak base resin, D301, D303 , D380, D301K macroporous weak alkali resin and one or more of ODS filler (i.e. octadecyl bonded silica gel); further preferably macroporous strong basic anion exchange resin DKx4 and/or macroporous Weakly basic anion exchange resins D303, D301K; more preferably DKx4 resin. .
  • the column chromatography in step 3) of the present invention uses an elution solvent to perform gradient elution on the chromatography column;
  • the elution solvent is preferably an acidic organic solvent and/or acid or alkaline water;
  • the step of gradient elution includes: removing impurities with 0.05%-2% (M/M) alkaline water, and eluting with 60%-80% (V/V) acidic ethanol with pH 2-5 Obtain the target product part.
  • the above gradient elution step not only makes the target product partly pure, but also regenerates the chromatographic column continuously and can be recycled.
  • the eluent obtained by concentration in step 3) of the present invention is that the relative density measured when the eluent is concentrated to 50° C. is 1.05 to 1.25, and is injected into the decolorization and desensitization tank while it is hot.
  • step 3) of the present invention further includes a step of recovering ethanol in the eluate.
  • the activated carbon described in step 3) of the present invention is in powder form or particles below 0.5 mm, imported or domestic activated carbon for decolorization and desensitization or its products.
  • the activated carbon input in the decolorization and desensitization tank described in step 3) of the present invention is 0.2-0.5% of the total CBD content (W/W). More preferably, the decolorization, desensitization and desensitization step in step 3) of the present invention is completed by stirring at 45-75°C for 0.5-1 h.
  • the step 3) of the present invention also includes filtration of the decolorized and desensitized CBD concentrate, and the decolorization and desensitization device is a 500-800 mesh titanium rod filter.
  • the step 3) is: the relative density measured when the elution section rich in CBD is concentrated to 50° C. is 1.05 ⁇ 1.25, then it is poured into the decolorization tank while it is hot, and the total content of CBD is added 0.2-0.5% (W/W) powdered activated carbon powder, stirred at 45-75°C for 0.5-1 h, and filtered through a titanium rod filter to obtain a decolorized and desensitized CBD stock solution.
  • the specific solvent mentioned in step 4) of the present invention refers to one or several multi-systems in different ratios among butane, pentane, hexane, heptane, ethyl acetate and ethanol. Further preferred solvents are hexane, heptane, ethyl acetate, acetone and ethanol. Ethanol here refers to an ethanol-water (or acetone-water) system, and the crystal concentration is 5-65% (V/V), more preferably 10-25%.
  • the supersaturated dissolution temperature of adding a specific solvent in step 4) of the present invention is 10-80°C, for example, it can be 10°C, 10.1°C, 10.5°C, 10.7°C, 11°C, 12°C, 15°C. °C ⁇ 18°C ⁇ 20°C ⁇ 23°C ⁇ 25°C ⁇ 26°C ⁇ 29°C ⁇ 30°C ⁇ 35°C ⁇ 37°C ⁇ 40°C ⁇ 40.5°C ⁇ 45°C ⁇ 50°C ⁇ 55°C ⁇ 60°C ⁇ 65°C ⁇ 70°C, 75°C, 80°C; more preferably 20 to 60°C; more preferably 30 to 50°C.
  • the step 4) is: concentrating the CBD decolorized and desensitized stock solution obtained in step 3) to a relative density of 1.05 to 1.35 when measured at 50°C, and measuring the ethanol in the eluate Recover, add n-hexane to supersaturate and dissolve at a temperature of 10-80°C, stir slowly, and incubate for 36 hours to obtain crystal particles.
  • the alcohol concentration for washing crystals in step 4) of the present invention is 0-40% (V/V), more preferably 5-15% (V/V).
  • the washing temperature in step 4) of the present invention is 0-24°C, for example, it can be 0°C, 0.1°C, 0.5°C, 0.8°C, 1.0°C, 1.5°C, 2.0°C, 2.5°C, 3°C, 4°C, 5°C, 6°C, 7.0°C, 8°C, 9°C, 10°C, 11°C, 13°C, 15°C, 17°C, 19°C, 20°C, 24°C, etc.; more preferably 5 -20°C; more preferably 10-15°C.
  • the step 4) is: the crystals are washed with purified water or 5-40% (V/V) ethanol at a temperature of 0-24° C. to obtain the initial product.
  • the drying method in step 4) of the present invention includes but is not limited to one or more of spray drying, vacuum drying, freeze drying, near infrared drying, and microwave drying.
  • the drying temperature in step 4) of the present invention is not more than 65°C.
  • the step 9) of the present invention also includes the step of pulverizing the obtained cannabidiol into powder; the pulverization method includes vapor stream pulverization and/or freezing pulverization; the temperature of the pulverized material is not Over 65°C.
  • a method for extracting cannabidiol from cannabis which includes the following steps:
  • the alcohol solution of the precipitate was subjected to column chromatography.
  • the column packing used in the column chromatography was strong base No. 4, strong base DKx4, Shanghai 763#, #702 weak base resin, D301, D303, D380, D301K,
  • One or more of macroporous weak base resins the specific steps include: first eluting with 0-60% (V/V) ethanol to remove impurities, and then eluting with 60-80% (V/V) ethanol Obtain the target product part, and finally eluted with 90-95% (V/V) ethanol to regenerate the chromatography column;
  • the eluent obtained by concentration is concentrated under reduced pressure to a density of 1.05 ⁇ 1.25 (alcohol content 40 ⁇ 70%), put into the decolorization tank, add activated carbon at 0.2 ⁇ 0.5% of the CBD content (W/W), stir at 45 ⁇ 75°C for 30min , Filter, get decolorization and desensitization solution for heat removal;
  • Step 4) the initial product is dried at a temperature of 40-55° C. and then pulverized to obtain a finished product of cannabidiol.
  • the descriptions of "4 to 8 times the amount of medicinal materials” or “1/10 to 1/100 times the amount of medicinal materials” in this application mean that the volume of the solvent used, such as ethanol or water, is 2 ⁇ 8 times or 1-10 times, for example, the medicinal powder is 1g, and the amount of extraction solvent ethanol is 2ml-8ml.
  • the present invention extracts cannabidiol (CBD) from hemp.
  • CBD cannabidiol
  • the process route is gradually optimized, so that the yield, purity and crystal form of cannabidiol (CBD) are significantly improved, especially the dynamic crystallization process It is suitable for large-scale production, and the physical properties of the obtained crystals are relatively better, which is conducive to subsequent formulation development; in addition, the final CBD raw material can meet the requirements of injection level, which extends the scope of its modern application.
  • the raw materials and reagents used in the method for extracting cannabidiol from hemp provided by the present invention can be purchased from the market.
  • the following examples further illustrate the present invention:
  • the example section provides that the method of the present invention is used to prepare cannabidiol under different technical parameters.
  • the amount of raw material industrial hemp flower, leaf, root, stalk core and/or seed meal used in each example is all 600 kg, no further explanation below.
  • the alcohol solution of the precipitate is subjected to column chromatography.
  • the packing of the chromatography column is a strong base DKx4, and the elution solvent is acidic ethanol and alkaline water.
  • the elution steps include: first use 2% alkaline water, pH 3.5 30% (V/V) ethanol to remove impurities, and then eluted with 80% (V/V) ethanol at pH 5 to obtain the target product part, and finally washed with 95% (V/V) ethanol at pH 5 Take off to regenerate the chromatography column;
  • step 7) Concentrate the eluent obtained in step 6) under reduced pressure to a density of 1.05 (65% alcohol content), put it into a decolorization tank, add activated carbon at 0.3% of the CBD content (W/W), stir at 45°C for 30 minutes, and filter. Obtain a decolorizing and desensitizing solution that removes heat;
  • step 8) Concentrate the decolorization and desensitization solution obtained in step 7) to a relative density of 1.25, recover the ethanol, add hexane-ethyl acetate (10:1, V/V) while hot to dissolve it oversaturated, and pour the clear solution into the crystallization tank , Incubate at -10°C for 12h, filter to obtain crystals;
  • step 9 Vacuum drying the initial product described in step 9), and pulverize it with steam flow to below 200 meshes to obtain cannabidiol, namely product 1.
  • step 7) Concentrate the eluate obtained in step 6) under reduced pressure and distill to a density of 1.12 (alcohol 55%), put it into the decolorization tank, add activated carbon at 0.5% of the CBD content (W/W), stir at 50°C for 30 minutes, and filter. Obtain a decolorizing and desensitizing solution that removes heat;
  • step 8) Concentrate the decolorization and desensitization solution obtained in step 7) to a relative density of 1.35, recover the ethanol, add heptane while it is hot to supersaturate the solution, transfer the clear solution into a crystallization tank, incubate at 10°C for 72 hours, and filter to obtain crystals;
  • step 9) Add 5% (V/V) ethanol to wash the crystals described in step 8) at a temperature of 24°C to obtain the initial product;
  • step 10) The initial product described in step 9) is dissolved in 30% ethanol-water, passed through a 1000D microporous membrane, and freeze-dried to obtain cannabidiol;
  • step 11 The cannabidiol obtained in step 10) is frozen and crushed to obtain product 2.
  • the elution steps include: First eluting with 60% (V/V) ethanol to remove impurities, then eluting with 75% (V/V) ethanol to obtain the target product part, and finally eluting with 100% (V/V) ethanol to make the layer Column regeneration;
  • step 7) Concentrate the eluent obtained in step 6) under reduced pressure to a density of 1.17 (46% alcohol content), put it into the decolorization tank, add activated carbon at 0.2% of the CBD content (W/W), stir at 75°C for 30 min, filter, Obtain a decolorizing and desensitizing solution that removes heat;
  • step 8) Concentrate the decolorization and desensitization solution obtained in step 7) to a relative density of 1.05, recover the ethanol, add 25% (V/V) ethanol while it is hot, and dissolve it in supersaturation at 70°C. The clear solution is poured into a crystallization tank. Incubate at 20°C for 48 hours and filter to obtain crystal 1;
  • step 9) Put the crystallized substance 1 obtained in step 8) into the crystallization tank again, add 25% (V/V) ethanol, dissolve in supersaturation at 70°C, incubate at 0°C for 48h, and filter to obtain the initial product of cannabidiol;
  • the alcohol solution of the precipitate is subjected to column chromatography.
  • the filler of the chromatography column is AB-8, and the elution solvent is ethanol and water.
  • the elution steps include: first wash with 30% (V/V) ethanol Removal of impurities, then eluting with 80% (V/V) ethanol to obtain the target product part, and finally eluting with 95% (V/V) ethanol to regenerate the chromatography column;
  • step 6) The relative density of the eluent obtained in step 6) is 1.15 when the concentration reaches 50°C, and it is dissolved in 100% (V/V) ethanol supersaturated at 10°C to obtain crystals;
  • step 8) Add purified water to wash the crystals described in step 7) at a temperature of 0°C to obtain the initial product;
  • Example 4 CBD transfer and content in finished products obtained by different preparation methods
  • the number of theoretical plates should not be less than 2500 based on the CBD peak.
  • Preparation of reference substance solution accurately weigh the CBD reference substance, add methanol (1:1) to make a reference substance solution containing 0.1mg per lml, and get it; accurately weigh the tetrahydrocannabinol reference substance, add methanol (1 :1) Prepare a reference solution containing 0.01mg per lml, and get it.
  • Preparation of solid test solution Take about 25mg of the CBD initial or finished product, accurately weigh it, place it in a 25ml measuring flask, add 20ml of acetonitrile-water (1:1), ultrasonically treat for 10 minutes, add acetonitrile-water (1: 1) Dilute to the mark, shake it, filter with a microporous membrane (0.45pm), take the filtrate, and get it.
  • Preparation of the liquid test solution take the liquid intermediate sample, place it in a 25ml measuring flask, dilute with acetonitrile-water (1:1) as appropriate, sonicate for 10 minutes, add acetonitrile-water (1:1) to the mark, Shake the hook, filter with a microporous membrane (0.45pm), take the filtrate, and get it.
  • Determination method accurately draw 10 ⁇ l each of the reference solution and the test solution, and inject it into the liquid chromatograph for determination.
  • the acid-adjusting water precipitation process can significantly increase the yield of CBD, and different solvents and crystallization times also affect the content and yield of CBD products to a certain extent.
  • Example 5 The influence of different crystallization solvents on crystallization
  • n-hexane and n-heptane are equivalent, so the list of n-heptane and its combined solvent with ethyl acetate will not be repeated in the table.
  • n-hexane and heptane and the combined solvent of the two and ethyl acetate have better crystallization effects, and the combined solvent of the two and ethyl acetate has a concentration ratio of 100:10.
  • the crystallization effect is the best. The reason is that the mother liquor dissolves the largest amount of impurities in the presence of ethyl acetate.
  • the size of crystal formation is related to the speed of stirring.
  • the data of different rotation speeds and crystal size are collected in a workshop 500L scraper crystallizer (equipment of different scales, the rotation speed and the blade shear force are quite different), and they are all dissolved in heptane: ethyl acetate (100:10), Stir and crystallize at 10°C. After finishing, filter out the crystals, mix well, and place a sample in the sample cell of the laser particle size analyzer.
  • Measurement conditions Omega LS-C (IIA) dry laser particle size analyzer, medium: air; medium refractive index: 1.00; sample refractive index: 1.70; shading ratio: 2.4%; cut-off lower limit: 0.20 ⁇ m, cut-off upper limit: 500.00 ⁇ m; Analysis mode: polydis.
  • Other measurement items are carried out according to the 2015 Chinese Pharmacopoeia.
  • the parameters of the powder are measured from each group of airflow crushed to 200 mesh crystal powder; the gravity flow rate is taken from the time when each group of 10g 200 mesh crystal powder completely flows out.
  • the rotation speed in dynamic crystallization affects crystal formation and growth. Slow to medium speed is appropriate, and the physical properties of the crystal are relatively better, which is conducive to subsequent formulation development; slower and static crystallization tends to agglomerate, and faster dominate affect the yield.
  • Preparation of test rabbits Take experimental robust rabbits of more than 1.5kg, no disease, pregnancy or medication records, unified management of boarding and lodging environment, especially breeding room temperature fluctuation should not exceed 3°C, and ensure the rectal temperature within 3-7 days before the experiment
  • the continuous measurement is stable (38.8 ⁇ 0.8°C), that is, the maximum and minimum temperature of the rectal temperature of each rabbit for a continuous week does not exceed 0.4°C, otherwise, it is unqualified and cannot be used as an experimental object.
  • the measured value is uniformly taken at 6cm inside the anus, and the value is stable for 2 minutes
  • the rabbits should stop feeding 2 hours before the test to avoid false positive effects on the experimental results.
  • Injection preparation All appliances that may come into contact with drugs or introduce external heat sources must be sterilized at 250°C for 30 minutes in advance. If it cannot be sterilized by high heat, it should be sterilized in advance by other suitable sterilization methods.
  • CBD crystal powder Take the finished CBD crystal powder, dissolve it with injection-grade soybean oil in an ultra-clean workbench, and configure it into a 0.1mg/ml injection, each with 2ml ampoules, and each batch of finished products will make 10 for use.
  • the total temperature of the rabbits exceeds 3.5°C; that is, the rabbit heat source test result is determined to be positive, and the batch of samples is unqualified.

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Abstract

本发明涉及医药技术领域，特别涉及大麻二酚(CBD)的制备方法，本发明制备方法包括提取、脱羧、酸水沉降、纯化、结晶等步骤，本发明中采用提取后转化脱羧可以使脱羧更完全；酸水沉极大提升大麻二酚(CBD)的得率；用大孔型离子交换树脂柱层析纯化，除杂效果更好；刮板式桨叶与特定溶媒组合的动态结晶工艺结晶效率更高，更适合大规模的生产，且所得晶体的物理性征相对更佳，利于后续制剂开发；通过本发明方法制备所得的CBD原料能够达到注射级别的要求，延伸了其现代化应用的范畴。

Description

从大麻中提取大麻二酚的方法
本申请要求于2019年04月30日提交中国专利局、申请号为201910363211.4、发明名称为“从大麻中提取大麻二酚的方法”的中国专利申请的优先权，其全部内容通过引用结合在本申请中。
技术领域
本发明涉及医药技术领域，特别涉及从大麻中提取大麻二酚的方法。
背景技术
工业大麻(学名：Cannabis sativa L.)为桑科、大麻属植物，生长期花叶中的四氢大麻酚(THC)含量小于千分之三，不具备提取四氢大麻酚的价值或直接作为毒品吸食，可以合法进行规模化种植与工业化开发利用，其经济、药用价值极高。
大麻酚类化合物是大麻植株中含有的一类活性物质，主要有四氢大麻酚(THC)、四氢次大麻酚(THCV)、大麻二酚(CBD)、大麻萜酚(CBG)、次大麻二酚(CBDV)等，这五者占大麻酚类化合物的90％以上。大麻二酚(CBD，cannabidiol)，为植株中最重要的非成瘾性成分之一，具有抗痉挛、抗风湿性关节炎、抗焦虑等药理活性，且能阻碍四氢大麻酚(THC)对人体神经系统的不良影响，成为药物开发的热点。
专利PCT/CN2017/071993，提及一种从大麻中提取大麻二酚的方法：即将大麻的提取部位粉碎、烘干炮制，得到药材粉末，采用30-100％(V/V)的乙醇提取，得到提取液，减压浓缩，得到浸膏，趁热加水进行水沉，去除水层杂质，在沉淀中加入10-100％(V/V)乙醇溶解，上柱层析，得到富含CBD的目标产物，而后，以乙醇等有机溶剂结晶，将晶体洗涤后干燥，即得大麻二酚成品，纯度在99％左右。
但随着生产规模的持续放大以及市场对CBD品质不断提升的要求，现工艺渐渐呈现出以下问题：
1)大麻花叶炮制脱羧环节，因其特别蓬松，放大生产后，对生产设备要求较高，极易成为限速环节且因放大效应，脱羧比例有不同程度的降 低，造成显著的批间和批内差异；
2)因花叶质量难以均一(比如老叶与新叶，不同地块土质等)，当水沉液中电解质成分(尚不明确来源和组成)较多时，易与大麻素成分形成小胶团，分散在水沉液中，随废水排走而造成大量的目标产物损耗；
3)花叶提取纯化后，结晶是在低温或常温常压下，静置12-72h进行的，由于放大效应，单次结晶不同结晶部位(如晶体表面、罐体侧壁和底部)纯度差异较大，混批后，含量已经达不到99％以上，需要重结晶；此外，这样形成的晶体通常为一大块且非常紧致，难以收获和粉碎，已然不适合规模化生产的要求。
4)通过以上工艺过程，我们可得纯度很高的成品，但做为注射级原料，其安全性仍不能得到保障，影响工业化应用。
因此，目前亟需提供一种可以大规模工业化制备且生产更高效，质量更安全大麻二酚的方法，以推动市场要求不断提升的大麻产业发展需求。
发明内容
有鉴于此，本发明提供一种从大麻中提取大麻二酚的方法。本发明提供了一种注射级大麻二酚原料的制备方法，以解决原工艺高损耗低效能等问题，并从根本上解决结晶工业化难题，同时，使得CBD成品可以达到注射级原料的要求。
为了实现上述发明目的，本发明提供以下技术方案：
本发明提供了从大麻中提取大麻二酚的方法，包括如下步骤：
步骤1：取大麻粉碎、烘干，获得大麻粉末；
步骤2：取大麻粉末经醇提后收集提取液、第一浓缩、与水混合后脱羧、再经第二浓缩得浸膏；所述脱羧的温度为60℃～130℃，时间为1h～3h；
步骤3：取所述浸膏与水混合，调节pH值为3.5～6.5，水沉，离心，收集沉淀，再与乙醇混合，纯化，经第三浓缩获得浓缩液；再经脱色脱敏，过滤，第四浓缩，获得稠膏；
步骤4：取所述稠膏与溶媒于10℃～80℃混合，持续搅拌于-10℃～20℃ 结晶12h～72h，洗涤，干燥。
在本发明的一些具体实施方案中，所述大麻为工业大麻、中间型大麻或药用大麻中的一种或两者以上的组合。
在本发明的一些具体实施方案中，所述大麻的提取部位为大麻花、大麻叶、大麻根、大麻秆芯或大麻籽粕中的一种或两种以上的组合。
在本发明的一些具体实施方案中，步骤1中所述粉碎至10～80目。
在本发明的一些具体实施方案中，步骤2中所述醇提采用30～100％(V/V)的乙醇提取，所述提取包括回流提取、超声提取和/或浸泡提取；所述醇提的次数为1～3次。
在本发明的一些具体实施方案中，所述的回流提取包括：采用2～10倍药材量的乙醇进行回流提取1～3次，每次0.5～3h；
所述的超声提取包括：采用2～10倍药材量的乙醇进行超声提取1～3次，每次0.1～1h；
所述的浸泡提取包括：采用2～10倍药材量的乙醇进行浸泡提取1～3次，每次0.5～5h。
在本发明的一些具体实施方案中，步骤2中所述第一浓缩或第二浓缩为50～70℃减压浓缩至相对密度为1.05～1.35(50℃测定值)。
在本发明的一些具体实施方案中，步骤2中所述水的加入量为药材1/10～1/100倍量(W/V)。
在本发明的一些具体实施方案中，所述调节pH值采用有机酸、无机酸或碱，可以包括甲酸，乙酸，草酸，盐酸，硝酸，硫酸等之中的一种或数种。更优选的是甲酸，草酸和盐酸。所述的碱为氢氧化钠，氢氧化钾或氨水，更优选的是氢氧化钠。
在本发明的一些具体实施方案中，步骤3中所述水的用量为1～10倍药材量，所述水沉的温度为0℃～20℃，所述水沉的时间为1～48h。
在本发明的一些具体实施方案中，所述离心的转速为4000rpm～10000rpm，所述离心的时间为3min～30min。
在本发明的一些具体实施方案中，步骤3中所述沉淀与乙醇的质量体积比为4:(1～4)；所述纯化为大孔型离子交换树脂的柱层析，所述柱层 析采用洗脱溶剂进行梯度洗脱；所述梯度洗脱为：用0.05％～2％(M/M)的碱水除杂，再用pH 2～5的60％～80％(V/V)的酸性乙醇洗脱获得洗脱液；所述第三浓缩为于50～70℃减压浓缩至相对密度为1.05～1.25(50℃测定值)，酒精度40％～70％。所述梯度洗脱用碱水为氢氧化钠，氢氧化钾或氨水的水溶液，更优选的是氢氧化钠水溶液
在本发明的一些具体实施方案中，层析柱的填料包括强碱4号，强碱DKx4、上海763 #、 #702弱碱树脂，D301、D303、D380、D301K、大孔弱碱树脂中的一种或多种。
在本发明的一些具体实施方案中，所述梯度洗脱的步骤包括先用0.05％～2％(M/M)的碱水除杂，再用pH 2～5的60％～80％(V/V)的酸性乙醇洗脱获得目标产物部分，最后用pH 2～5的90-95％(V/V)的酸性乙醇冲柱以使层析柱再生。上述梯度洗脱的步骤不仅使得目标产物部分纯度高，色泽好而且使层析柱不断再生，可循环使用。
在本发明的一些具体实施方案中，步骤3中所述脱色脱敏采用活性炭，所述活性炭的加入量为所述浓缩液中大麻二酚质量的0.2％～0.5％(W/W)，所述脱色脱敏的温度为45℃～75℃，所述脱色脱敏的时间为0.5h～1h；所述过滤为压滤或抽滤，所述过滤采用的滤网孔径不小于400目。
在本发明的一些具体实施方案中，所述活性炭为粉末状或0.05mm～0.5mm的颗粒。
在本发明的一些具体实施方案中，步骤4中所述溶媒为丁烷、戊烷、己烷、庚烷、乙酸乙酯、丙酮或乙醇中的一种或两者以上的组合；所述洗涤于0℃～24℃采用水或5％～40％(V/V)乙醇洗涤；所述干燥包括喷雾干燥、真空干燥、冷冻干燥、近红外干燥或微波干燥中的一种或多种，所述干燥的温度为30℃～65℃。
作为优选，所述干燥的温度为40℃～55℃。
在本发明的一些具体实施方案中，在干燥之后还包括粉碎的步骤，所述粉碎包括汽流粉碎和/或冷冻粉碎，所述粉碎的温度为0℃～65℃。
在本发明的一些具体实施方案中，所述溶媒为己烷、庚烷、乙酸乙酯、 丙酮或乙醇中的一种或两者以上的组合。此处的乙醇(或丙酮)指的是乙醇-水(或丙酮-水)系统，结晶浓度为5％～65％(V/V)，更优选为10％～25％(V/V)。
本发明从大麻中提取大麻二酚(CBD)，在持续放大中，全面优化工艺路线：1)提取后转化脱羧可以使脱羧更完全(见图6)；2)酸水沉极大提升大麻二酚(CBD)的得率；3)采用大孔型离子交换树脂柱层析，除杂效果更好4)刮板式桨叶与特定溶媒组合的动态结晶工艺时结晶效率更高，适合大规模的生产(见图7)，且所得晶体的物理性征相对更佳，利于后续制剂开发；5)采用本发明方法制备得的CBD原料能够达到注射级别的要求，延伸了其现代化应用的范畴。
附图说明
为了更清楚地说明本发明实施例或现有技术中的技术方案，下面将对实施例或现有技术描述中所需要使用的附图作简单地介绍。
图1示CBD标准品(sigma)的色谱图；
图2示CBD产物1的色谱图；
图3示CBD产物2的色谱图；
图4示CBD产物3的色谱图；
图5示CBD产物4的色谱图；
图6示上样液的的色谱图；其中，图6(A)示对比例制得的产物4(先脱羧后提取)，图6(B)示本发明实施例1制得的产物1(先提取后脱羧)；
图7示结晶样品图；其中，图7(A)～图7(B)示本发明实施例1的结晶样品；图7(C)～图7(E)为对比例的结晶样品。
具体实施方式
本发明公开了一种从大麻中提取大麻二酚的方法，本领域技术人员可以借鉴本文内容，适当改进工艺参数实现。特别需要指出的是，所有类似的替换和改动对本领域技术人员来说是显而易见的，它们都被视为包括在本发明。本发明的方法及应用已经通过较佳实施例进行了描述，相关人员 明显能在不脱离本发明内容、精神和范围内对本文所述的方法和应用进行改动或适当变更与组合，来实现和应用本发明技术。
本发明旨在提供一种注射级大麻二酚原料的制备方法，以解决原工艺高损耗低效能等问题，同时，把CBD成品品质推上注射级的高度。
为了实现上述目的，本发明提供了一种从大麻中提取大麻二酚的方法，该方法包括以下步骤：
1)将大麻提取部位粉碎，得到药材粉末；
2)将药材粉末采用30～100％(V/V)的乙醇提取，得到提取液；
将提取液减压浓缩至无醇，回收乙醇，加入药材1/10～1/100倍量(W/V)的纯化水，脱羧，继续常压浓缩1～2h，得到浸膏；所述脱羧的温度为60℃～130℃，时间为1h～3h；
3)将浸膏以纯化水分散，以有机酸或无机酸调节pH为3.5～6.5，去除水溶性杂质同时，使CBD最大程度沉降，得到水沉液；
将水沉液离心，在离心获得的沉淀中加入10～100％(V/V)乙醇溶解，得到沉淀物的醇溶液；将沉淀物的醇溶液进行柱层析；
浓缩得到的目标段洗脱液，减压浓缩至密度1.05～1.25(酒精度40～70％)，打入脱色罐，按CBD总含量(W/W)的0.2～0.5％加入活性炭，45～75℃搅拌0.5～1h，过滤，得除热源的脱色脱敏液；
4)浓缩步骤7)的脱色脱敏液至无水稠膏，加入特定的溶媒过饱和溶解，将上清液置于结晶罐中，低温至常温，持续搅拌结晶12～72h，最终获得颗粒大小均一结晶物；
将结晶物，加入纯化水或乙醇洗涤，得到初品；
将所得初品用适宜的干燥方法干燥，粉碎，即得大麻二酚成品。
优选的，本发明所述的大麻选自工业大麻、中间型大麻或药用大麻中的一种或两种以上的组合。
优选的，本发明所述的步骤1)中提取部位选自大麻花、大麻叶、大麻根、大麻秆芯和大麻籽粕中的一种或两种以上的组合。优选的，本发明所述的步骤1)中提取部位为大麻花、大麻叶。
优选的，本发明所述的步骤1)中所述大麻提取部位粉碎至10～80目， 例如可以是10目、12目、14目、16目、18目、20目、25目、30目、35目、40目、45目、50目、60目、70目、80目等等；进一步优选为20～60目；更优选为25-50目；最优选为40目。在本发明的一个典型实施方式中，所述的步骤1)中将大麻花、叶粉碎至前述目数范围内，可以使得大麻二酚在后续的乙醇提取步骤中充分提取出来。
优选的，本发明所述的步骤2)中所述乙醇的用量为4-10倍药材量，例如可以是4倍、4.5倍、5倍、6倍、7倍、8倍、9倍、10倍等药材量；进一步优选为4～8倍药材量；更优选为4-6倍药材量。
优选的，本发明所述的步骤2)中所述乙醇提取的次数为1～3次。
优选的，本发明所述的步骤2)中所述乙醇提取的提取方式为回流提取、超声提取和/或浸泡提取。
进一步优选的，所述回流提取的时间为每次提取0.5～3h；所述超声提取的时间为每次提取0.1～lh；所述浸泡提取的时间为每次提取0.5～5h。
在本发明的一个典型实施方式中，所述步骤2)为：将药材粉末采用4～8倍药材量的30～100％(V/V)的乙醇提取1～3次，得到提取液。
优选的，本发明所述的步骤2)中所述将提取液浓缩为浓缩至50℃时测量的相对密度为1.05～1.35。
优选的，本发明所述的步骤3)中所述水沉的用水量为1～10倍药材量，例如可以是1倍、1.1倍、1.2倍、1.3倍、1.4倍、1.5倍、1.6倍、1.7倍、1.8倍、1.9倍、2倍、2.25倍、2.5倍、3倍、3.5倍、4倍、4.5倍、5倍、5.5倍、6倍、6.5倍、7倍、7.5倍、8倍、8.5倍、9倍、9.5倍、10倍药材量；进一步优选为2-8倍药材量；更优选为3-6倍药材量。
优选的，本发明所述的步骤3)中所述水沉调节pH的酸为甲酸，乙酸，草酸，盐酸，硝酸，硫酸等之中的一种或数种。更优选的是甲酸，草酸和盐酸。所述的碱为氢氧化钠，氢氧化钾或氨水，更优选的是氢氧化钠。优选的，本发明所述的步骤3)中所述水沉调节pH为2.5～6.5，例如可以是2.5、2.7、2.9、3.0、3.3、3.5、3.7、3.9、4.0、4.5、5.0、5.5、6.0、6.5等，更优选为3.5～5.5。
优选的，本发明所述的步骤3)中所述水沉的温度为0～20℃，例如 可以是0℃、0.1℃、0.5℃、0.8℃、1.0℃、1.5℃、1.75℃、2℃、3℃、4℃、5℃、6℃、7℃、8℃、9℃、10℃、11℃、13℃、15℃、17℃、18℃、19℃、20℃等；进一步优选为2～18℃；更优选为5～15℃。
优选的，本发明所述的步骤3)中所述水沉的时间为l-48h，例如可以是lh、l.lh、1.3h、1.5h、1.9h、2h、2.5h、3h、4h、5h、6h、7h、8h、9h、10h、15h、20h、30h、40h、48h等；进一步优选为2-36h；更优选为5-30h。
在本发明的一个典型实施方式中，所述步骤3)为：将浸膏用其1～10倍药材量的纯化水在0～20℃的温度下进行水沉l～48h，去除杂质，得到水沉液。
优选的，本发明所述的步骤3)中所述柱层析所用的层析柱的填料包括但不限于强碱4号，强碱DKx4、上海763#、#702弱碱树脂，D301、D303、D380、D301K大孔弱碱树脂以及ODS填料(即十八烷基键合硅胶)中的一种或多种，；进一步优选为大孔型强碱性阴离子交换树脂DKx4和/或大孔型弱碱性阴离子交换树脂D303、D301K；更优选为DKx4树脂。。
优选的，本发明所述的步骤3)中所述柱层析为采用洗脱溶剂对层析柱进行梯度洗脱；所述洗脱溶剂优选为酸性有机溶剂和/或酸、碱水；进一步优选的，所述梯度洗脱的步骤包括：0.05％～2％(M/M)的碱水除杂，再用pH 2～5的60％～80％(V/V)的酸性乙醇洗脱获得目标产物部分。上述梯度洗脱的步骤不仅使得目标产物部分纯度高，而且使层析柱不断再生，可循环使用。
优选的，本发明所述的步骤3)中所述浓缩得到的洗脱液为将洗脱液浓缩至50℃时测量的相对密度为1.05～1.25，趁热打入脱色脱敏罐。进一步优选的，本发明所述的步骤3)中还包括对洗脱液中乙醇的回收步骤。
优选的，本发明所述的步骤3)中所述的活性炭为粉末状或0.5mm以下的颗粒，进口或国产的脱色脱敏用活性炭或其制品。
优选的，本发明所述的步骤3)中所述的脱色脱敏罐中活性炭投入量为CBD总含量(W/W)的0.2～0.5％。进一步优选的，本发明所述的步骤3)中脱色脱敏脱敏步骤是在45～75℃下搅拌0.5～1h完成的。本发明所述的步骤3)中还包括对脱色脱敏脱敏的CBD浓缩液的过滤，脱色脱敏装 置为500～800目的钛棒滤器。
在本发明的一个典型实施方式中，所述步骤3)为：将富含CBD的洗脱段浓缩至50℃时测量的相对密度为1.05～1.25，趁热打入脱色罐，加入CBD总含量0.2～0.5％(W/W)的粉末状活性炭粉，45～75℃下搅拌0.5～1h后，通过钛棒滤器压滤，得脱色脱敏的CBD原液。
优选的，本发明所述的步骤4)中所述的特定溶媒指的是丁烷，戊烷，己烷，庚烷，乙酸乙酯及乙醇中的一种或几种不同比例的多元体系。进一步优选的溶媒是己烷、庚烷、乙酸乙酯、丙酮和乙醇。此处的乙醇指的是乙醇-水(或丙酮-水)系统，结晶浓度为5～65％(V/V)，更优选为10～25％。
优选的，本发明所述的步骤4)中所述加入特定溶媒过饱和溶解的温度为10～80℃，例如可以是10℃、10.1℃、10.5℃、10.7℃、11℃、12℃、15℃、18℃、20℃、23℃、25℃、26℃、29℃、30℃、35℃、37℃、40℃、40.5℃、45℃、50℃、55℃、60℃、65℃、70℃、75℃、80℃；进一步优选为20～60℃；更优选为30～50℃。
在本发明的一个优选实施方式中，所述步骤4)为：浓缩步骤3)得到的CBD脱色脱敏的原液至50℃时测量的相对密度为1.05～1.35，对洗脱液中的乙醇进行回收，并在10～80℃的温度下加入正己烷过饱和溶解，慢速搅拌，孵育36h获得结晶物颗粒。
优选的，本发明所述的步骤4)中所述洗涤晶体的酒精浓度为0～40％(V/V)，更优选为5～15％(V/V)。
优选的，本发明所述的步骤4)中所述洗涤的温度为0～24℃，例如可以是0℃、0.1℃、0.5℃、0.8℃、1.0℃、1.5℃、2.0℃,2.5℃、3℃、4℃、5℃、6℃、7.0℃、8℃、9℃、10℃、11℃、13℃、15℃、17℃、19℃、20℃、24℃等；进一步优选为5～20℃；更优选为10～15℃。
在本发明的一个典型实施方式中，所述步骤4)为：结晶物在0～24℃的温度下，加入纯化水或5～40％(V/V)的乙醇洗涤，得到初品。
优选的，本发明所述的步骤4)中所述干燥的方式包括但不限于喷雾干燥、真空干燥、冷冻干燥、近红外干燥、微波干燥中的一种或几种。优选的，本发明所述的步骤4)中所述干燥的温度为不超过65℃。
进一步优选的，本发明所述的步骤9)中还包括将获得的大麻二酚粉碎成粉的步骤；所述粉碎的方式包括汽流粉碎、和/或冷冻粉碎；所述粉碎的物料温度不超过65℃。
在本发明的一个优选实施方式中，提供了一种从大麻中提取大麻二酚的方法，包括以下步骤：
1)将大麻的花叶粉碎至10～80目，得到药材粉末；
2)将药材粉末采用4～10倍药材量的30～100％(V/V)的乙醇提取1～3次，得到提取液；
将提取液减压浓缩至无醇味，50℃时测量的相对密度为1.05～1.35，加入药材量1/20倍量的纯化水，常压反应1～3h得到流膏；
3)将此流膏用其1～10倍药材量的纯化水分散，加酸调节pH为3.5～6.5，在0～20℃的温度下进行水沉l～48h，去除杂质，得到水沉液；
将水沉液离心，在离心获得的沉淀中加入10～100％(V/V)乙醇溶解，得到沉淀物的醇溶液；
将沉淀物的醇溶液进行柱层析，所述柱层析所用的层析柱填料为强碱4号，强碱DKx4、上海763#、#702弱碱树脂，D301、D303、D380、D301K、大孔弱碱树脂中的一种或多种，具体步骤包括：先用0～60％(V/V)的乙醇洗脱除杂，再用60～80％(V/V)的乙醇洗脱获得目标产物部分，最后用90～95％(V/V)的乙醇洗脱以使层析柱再生；
浓缩得到的洗脱液减压浓缩至密度1.05～1.25(酒精度40～70％)，打入脱色罐，按CBD含量(W/W)的0.2～0.5％加入活性炭，45～75℃搅拌30min，过滤，得除热源的脱色脱敏液；
4)将得到的洗脱液减压浓缩至50℃时测量的相对密度为1.05～1.35，对洗脱液中乙醇的进行回收，10～80℃的温度下加入某种特定溶媒过饱和溶解，清液置于结晶罐，-10～20℃孵育12～72h，过滤，获得结晶物；
将结晶物，在0～24℃的温度下，加入纯化水或5～40％(V/V)的乙醇洗涤，得到初品；
将步骤4)初品，在40～55℃的温度下干燥后粉碎，即得大麻二酚成品。
需要说明的是，本申请中的“4～8倍药材量”或“1/10～1/100倍药材量”等描述，是指采用的溶剂如乙醇或水的体积是药材质量的2～8倍或1～10倍，比如，药材粉末为1g，提取溶剂乙醇的用量为2ml～8ml。
本发明从大麻中提取大麻二酚(CBD)，在原工艺的持续放大中，逐渐优化工艺路线，使得大麻二酚(CBD)的得率、纯度、晶型均有明显提高，尤其动态结晶工艺更适合大规模的生产、且所得晶体的物理性征相对更佳，利于后续制剂开发；此外，最终所得的CBD原料能够达到注射级别的要求，延伸了其现代化应用的范畴。
本发明提供的从大麻中提取大麻二酚的方法中所用原料及试剂均可由市场购得。下面结合实施例，进一步阐述本发明：
实施例1大麻二酚的制备
实施例部分提供了采用本发明所述方法，在不同技术参数条件下，制备大麻二酚，各实施例中采用的原料工业大麻花、叶、根、秆芯和/或籽粕的量均为600千克，以下不再另做说明。
1)将工业大麻的整株粉碎，过80目筛，得到药材粉末，测得其水分含量为13.6％；
2)将上述药材粉末采用其4倍量、30％(V/V)的乙醇回流提取3次，每次0.5h，得到提取液；
3)将上述提取液(-0.08Mpa，60℃)减压浓缩至50℃时测量的相对密度为1.05，加入药材1/10倍量纯化水，常压浓缩1.5h(94℃)，得到流膏；
4)将上述流膏用其1倍量的纯化水分散，以冰醋酸调pH为5.5，在0℃的温度下水沉lh，去除杂质，得到水沉液；
5)将上述水沉液离心，转速5000转，在离心获得的沉淀中加入100％(V/V)乙醇溶解，得到沉淀物的醇溶液；
6)将沉淀物的醇溶液进行柱层析，其层析柱的填料为强碱DKx4，洗脱溶剂为酸性乙醇和碱水，洗脱的步骤包括：先依次用2％碱水、pH 3.5的30％(V/V)乙醇洗脱除杂，再用pH 5的80％(V/V)的乙醇洗脱获得目标 产物部分，最后用pH 5的95％(V/V)的乙醇洗脱以使层析柱再生；
7)浓缩步骤6)得到的洗脱液减压蒸馏至密度1.05(酒精度65％)，打入脱色罐，按CBD含量(W/W)的0.3％加入活性炭，45℃搅拌30min，过滤，得除热源的脱色脱敏液；
8)将步骤7)得到的脱色脱敏液浓缩至相对密度为1.25，乙醇回收，趁热加入己烷-乙酸乙酯(10:1，V/V)过饱和溶解，清液打入结晶罐，-10℃孵育12h，过滤，获得结晶物；
9)将步骤8)所述结晶物，在0℃下，加入纯化水洗涤，得到初品；
10)将步骤9)所述初品真空干燥，汽流粉碎至200目以下，得大麻二酚，即产物1。
实施例2:大麻二酚的制备
1)将工业大麻的花叶粉碎，过40目筛，得到药材粉末，测得其水分含量为14.2％；
2)将上述药材粉末采用其5倍量、70％(V/V)的乙醇超声辅助提取2次，每次0.5h，得到提取液；
3)将上述提取液，(-0.80Mpa，65℃)减压浓缩至50℃时测量的相对密度为1.35，加入药材1/20倍量纯化水，常压浓缩1h(94℃)，得到浸膏；
4)将上述浸膏用其10倍量的纯化水分散，以盐酸(10mol/L)调pH为2.5，在20℃的温度下水沉4h，去除杂质，得到水沉液；
5)将上述水沉液离心，转速10000转，在离心获得的沉淀中加入10％(V/V)乙醇溶解，得到沉淀物的醇溶液；
6)将沉淀物的醇溶液板框过滤后，进行柱层析，其层析柱的填料为D301K弱碱性阴离子交换树脂，洗脱溶剂为酸性乙醇和碱水，洗脱的步骤包括：先用0.5％碱水、pH 5的40％(V/V)的乙醇洗脱除杂，再用pH 2的60％(V/V)的乙醇洗脱获得目标产物部分，最后用pH 2的90％(V/V)的乙醇洗脱以使层析柱再生；
7)浓缩步骤6)得到的洗脱液减压蒸馏至密度1.12(酒精度55％)， 打入脱色罐，按CBD含量(W/W)的0.5％加入活性炭，50℃搅拌30min，过滤，得除热源的脱色脱敏液；
8)将步骤7)得到的脱色脱敏液浓缩至相对密度为1.35，乙醇回收，趁热加入庚烷过饱和溶解，清液打入结晶罐，10℃孵育72h，过滤，获得结晶物；
9)将步骤8)所述结晶物，在24℃的温度下，加入5％(V/V)乙醇洗涤，得到初品；
10)将步骤9)所述初品用30％乙醇-水溶解，通过1000D的微孔滤膜，冷冻干燥，即得大麻二酚；
11)冷冻粉碎步骤10)所得大麻二酚，即得产物2。
实施例3:大麻二酚的制备
1)将工业大麻的花叶粉碎，过10目筛，得到药材粉末，测得其水分含量为15.1％。
2)将上述药材粉末采用其10倍量、100％(V/V)的乙醇浸渍提取2次，每次2.5h，得到提取液；
3)将上述提取液，(-0.80Mpa，65℃)减压浓缩至50℃时测量相对密度为1.2,加入药材1/100倍量纯化水，常压煮沸(94℃)1.5h，得到浸膏；
4)将上述浸膏用其5倍量的纯化水分散，以硫酸(18mol/L)调pH为6.0，在10℃的温度下水沉18h，去除杂质，得到水沉液；
5)将上述水沉液离心，转速7500转，在离心获得的沉淀中加入60％(V/V)乙醇溶解，得到沉淀物的醇溶液；
6)将沉淀物的醇溶液10000转离心后，取上清进行柱层析，其层析柱的填料为ODS，采用中压至高压，洗脱溶剂为乙醇和水，洗脱的步骤包括：先用60％(V/V)的乙醇洗脱除杂，再用75％(V/V)的乙醇洗脱获得目标产物部分，最后用100％(V/V)的乙醇洗脱以使层析柱再生；
7)浓缩步骤6)得到的洗脱液减压蒸馏至密度1.17(酒精度46％)，打入脱色罐，按CBD含量(W/W)的0.2％加入活性炭，75℃搅拌30min， 过滤，得除热源的脱色脱敏液；
8)将步骤7)得到的脱色脱敏液浓缩至相对密度为1.05，乙醇回收，趁热加入25％(V/V)的乙醇，在70℃下过饱和溶解，清液打入结晶罐，20℃孵育48h，过滤，获得结晶物1；
9)将步骤8)得到的结晶物1再次投入结晶罐，加入25％(V/V)的乙醇，在70℃下过饱和溶解，0℃孵育48h，过滤，获得大麻二酚初品；
10)将步骤9)所述结晶物2真空干燥，即得大麻二酚；
11)汽流粉碎步骤9)所得大麻二酚至100目以下，即得产物3。
对比例:按照专利PCT/CN2017/071993所述方法制备的大麻二酚
1)将600kg工业大麻的花叶粉碎，过80目筛，在150℃的温度下烘干2h，得到药材粉末，测得其水分含量为4％；
2)将上述药材粉末采用其2倍量、30％(V/V)的乙醇回流提取3次，每次0.5h，得到提取液；
3)将上述提取液浓缩至50℃时测量的相对密度为1.05，得到浸膏；
4)将上述浸膏采用其1倍量的纯化水，在20℃的温度下水沉lh，去除杂质，得到水沉液；
5)将上述水沉液离心，转速5000转，在离心获得的沉淀中加入100％(V/V)乙醇溶解，得到沉淀物的醇溶液；
6)将沉淀物的醇溶液进行柱层析，其层析柱的填料为AB-8，洗脱溶剂为乙醇和水，洗脱的步骤包括：先用30％(V/V)的乙醇洗脱除杂，再用80％(V/V)的乙醇洗脱获得目标产物部分，最后用95％(V/V)的乙醇洗脱以使层析柱再生；
7)浓缩步骤6)得到的洗脱液至50℃时的相对密度为1.15，在10℃的温度下，用100％(V/V)乙醇过饱和溶解，获得结晶物；
8)将步骤7)所述结晶物，在0℃的温度下，加入纯化水洗涤，得到初品；
9)将步骤8)所述初品50℃，真空干燥，即得大麻二酚；
10)汽流粉碎步骤9)所得大麻二酚至100-200目，即得产物4。
实施例4:不同制法所得成品中CBD转移及含量情况
检测方法：
色谱条件与系统适用性试验：以十八烷基硅烷键合硅胶为填充剂；以乙腈为流动相A，以水为流动相B，按A(％):B(％)＝70:30进行等度洗脱；检测波长为210nm。理论板数按CBD峰计算应不低于2500。
对照品溶液的制备：精密称取CBD对照品，加甲醇(1:1)制成每lml各含0.1mg的对照品溶液，即得；精密称取四氢大麻酚对照品，加甲醇(1:1)制成每lml各含0.01mg的对照品溶液，即得。
固体供试品溶液的制备：取CBD初品或成品约25mg，精密称定，置25ml量瓶中，加乙腈-水(1:1)20ml，超声处理10分钟，加乙腈-水(1:1)稀释至刻度，摇勾，用微孔滤膜(0.45pm)滤过，取续滤液，即得。
液体供试品溶液的制备：取液态中间体样品，置25ml量瓶中，酌情加乙腈-水(1:1)稀释，超声处理10分钟，加乙腈-水(1:1)稀释至刻度，摇勾，用微孔滤膜(0.45pm)滤过，取续滤液，即得。
测定法：分别精密吸取对照品溶液与供试品溶液各10μl,注入液相色谱仪，测定，即得。
按照实施例1～3制备得到的产物的CBD转移及含量检测结果参见表1及图1～5。
表1.不同制法所得产物其CBD转移及含量检出情况
Figure PCTCN2020079693-appb-000001
注：药材(CBD+0.90CBDA)总含量为6.45‰。
综上，调酸水沉工艺能够显著提升CBD产率，不同溶剂和结晶 次数也在一定程度上影响了CBD产品的含量和收率。
实施例5:不同结晶溶剂对结晶影响
分别以石油醚、正己烷(与正庚烷等效)、正己烷：乙酸乙酯(100:5,10,20)、25％丙酮、25％乙醇，按照以上动态结晶条件进行8组试验，每组平行3次小实验，结果取平均值。组合溶剂(V/V)，配置或回收重复利用，均以密度计辅助较验。最终，从析晶时间(肉眼可见大量晶核)、晶体性征、收率、含量四个方面综合考察溶剂因素对结晶的影响。
表2.不同结晶溶剂对结晶影响
Figure PCTCN2020079693-appb-000002
注：正己烷与正庚烷等效，故表中不再重复罗列正庚烷及其与乙酸乙酯的组合溶剂。
不同溶剂有不同的结晶行为，其中，以正己烷和庚烷，及二者分别与乙酸乙酯的组合溶剂结晶效果较好，并且二者分别与乙酸乙酯的组合溶剂在浓度比100:10左右时，结晶效果最好，其原因是母液在乙酸乙酯存在时，溶解了最大量的杂质。
实施例6:动态结晶转速—晶体粒度表
动态结晶时，晶体生成的大小性征与搅拌的速度相关。不同转速-晶体粒度数据以车间500L刮板式结晶罐为例采集(不同规模的设备，转速与其桨叶剪切力有较大差异)，均以庚烷：乙酸乙酯(100:10)溶解，在10℃搅拌结晶，结束后，滤出晶体，混匀后，取样置于激光粒度分析仪样品池。
测定条件：欧美克LS-C(IIA)型干法激光粒度分析仪，介质：空气；介质折射率：1.00；样品折射率：1.70；遮光比：2.4％；截断下限：0.20μm，截断上限：500.00μm；分析模式：polydis.其他测定项参照2015版中国药典进行。
测定结果：
表3.不同搅拌速度对结晶性征的影响
Figure PCTCN2020079693-appb-000003
Figure PCTCN2020079693-appb-000004
注：粉体的参数测定，均取自各组气流粉碎至200目的晶粉；重力流出速度，取自各组10g 200目晶粉完全流出的时间。
综上，动态结晶中转速大小影响结晶形成和生长，以慢速至中速为宜，且晶体的物理性征相对更佳，利于后续制剂开发；较慢及静态结晶易结块，较快则严重影响收率。
实施例7:家兔注射CBD针的热源检查
供试家兔准备：取1.5kg以上实验用健壮家兔，无病、孕或给药记录，统一食宿环境管理，尤其饲育室温浮动不得超过3℃，并保证实验前3-7日内肛温连续测定稳定(38.8±0.8℃)，即每只家兔连续一周的肛温测定最高与最低温不超过0.4℃，否则，即为不合格，不得作为实验对象。测值统一取肛门内6cm处，稳定2min后的数值
家兔受试前2h时，停止供食，以避免对实验结果产生假阳性影响。
注射液制备：所有可能接触药品或引入外源热源的器具，均需提前置于250℃ 30min高热灭菌。不能高热灭菌的，应采用其他适宜灭菌方式提前灭菌。
取CBD成品晶粉,在超净工作台内，以注射级大豆油溶解，配置成为0.1mg/ml针剂，每支安瓿瓶2ml，每批次成品各制10支备用。
注射实验与统计：随机取3只合格受试家兔，测试初始肛温稳定无误后，15min内，对家兔耳静脉缓缓注射2ml提前预热至38℃的CBD注射液，每隔30min统一于肛门内6cm处(稳定2min)测定一次肛温，在连续6次采集结果中，如有任何一只家兔出现升温超过正常肛温0.6℃及其以上，或者3只升温均低于0.6℃但升温总和超过1.3℃，则需另取5只合格家兔给药复核。
若3只受试家兔中，有一只以上，肛温超过0.6℃及其以上；或在复试5只家兔中，有一只以上，肛温超过0.6℃及其以上；或总计8只受试家兔升温的总和超过3.5℃；即判定家兔热源测试结果阳性，该批样品不合格。
表4.家兔注射CBD后，体温统计情况
Figure PCTCN2020079693-appb-000005
以上所述仅是本发明的优选实施方式，应当指出，对于本技术领域的普通技术人员来说，在不脱离本发明原理的前提下，还可以做出若干改进和润饰，这些改进和润饰也应视为本发明的保护范围。

Claims (10)

  1. 从大麻中提取大麻二酚的方法，其特征在于，包括如下步骤：
    步骤1：取大麻粉碎、干燥，获得大麻粉末；
    步骤2：取大麻粉末经醇提后收集提取液、第一浓缩、与水混合后脱羧、再经第二浓缩得浸膏；所述脱羧的温度为60℃～130℃，时间为1h～3h；
    步骤3：取所述浸膏与水混合，调节pH值为3.5～6.5，水沉，离心，收集沉淀，再与乙醇混合，纯化，经第三浓缩获得浓缩液；再经脱色脱敏，过滤，第四浓缩，获得稠膏；
    步骤4：取所述稠膏与溶媒于10℃～80℃混合，持续搅拌于-10℃～20℃结晶12h～72h，洗涤，干燥。
  2. 如权利要求1所述的方法，其特征在于，所述大麻为工业大麻、中间型大麻或药用大麻中的一种或两者以上的组合；
    所述大麻的提取部位为大麻花、大麻叶、大麻根、大麻秆芯或大麻籽粕中的一种或两种以上的组合。
  3. 如权利要求1或2所述的方法，其特征在于，步骤1中所述粉碎至10～80目。
  4. 如权利要求1至3任一项所述的方法，其特征在于，步骤2中所述醇提采用30～100％(V/V)的乙醇提取，所述提取包括回流提取、超声提取和/或浸泡提取；所述醇提的次数为1～3次；
    所述的回流提取包括：采用2～10倍药材量的乙醇进行回流提取1～3次，每次0.5～3h；
    所述的超声提取包括：采用2～10倍药材量的乙醇进行超声提取1～3次，每次0.1～1h；
    所述的浸泡提取包括：采用2～10倍药材量的乙醇进行浸泡提取1～3次，每次0.5～5h。
  5. 如权利要求1至4任一项所述的方法，其特征在于，步骤2中所述第一浓缩或第二浓缩为50-70℃减压浓缩至相对密度为1.05～1.35(50℃ 测定值)。
  6. 如权利要求1至5任一项所述的方法，其特征在于，步骤2中所述水的加入量为药材1/10～1/100倍量(W/V)。
  7. 如权利要求1至6任一项所述的方法，其特征在于，步骤3中所述水的用量为1～10倍药材量，所述水沉的温度为0℃～20℃，所述水沉的时间为1～48h；
    所述离心的转速为4000rpm～10000rpm，所述离心的时间为3min～30min。
  8. 如权利要求1至7任一项所述的方法，其特征在于，步骤3所述纯化为装填大孔型离子交换树脂的柱层析，所述柱层析采用洗脱溶剂进行梯度洗脱；所述梯度洗脱为：用0.05％～2％(M/M)的碱水除杂，再用pH2～5的60％～80％(V/V)的酸性乙醇洗脱获得洗脱液；所述第三浓缩为于50～70℃减压浓缩至相对密度为1.05～1.25(50℃测定值)，酒精度40％～70％。
  9. 如权利要求1至8任一项所述的方法，其特征在于，步骤3中所述脱色脱敏采用活性炭，所述活性炭的加入量为所述浓缩液中大麻二酚质量的0.2％～0.5％(W/W)，所述脱色脱敏的温度为45℃～75℃，所述脱色脱敏的时间为0.5h～1h；所述过滤为压滤或抽滤，所述过滤采用的滤网孔径不小于400目。
  10. 如权利要求1至9任一项所述的方法，其特征在于，步骤4中所述溶媒为丁烷、戊烷、己烷、庚烷、乙酸乙酯、丙酮或乙醇中的一种或两者以上的组合；所述洗涤于0℃～24℃采用水或5％～40％(V/V)乙醇洗涤；所述干燥包括喷雾干燥、真空干燥、冷冻干燥、近红外干燥或微波干燥中的一种或多种，所述干燥的温度为30℃～65℃。
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