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WO2022115973A1 - Procédés de commutation de solvant pour la précipitation, le traitement et la purification de cannabinoïdes sélectionnés à partir de mélanges complexes concentrés - Google Patents

Procédés de commutation de solvant pour la précipitation, le traitement et la purification de cannabinoïdes sélectionnés à partir de mélanges complexes concentrés Download PDF

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WO2022115973A1
WO2022115973A1 PCT/CA2021/051745 CA2021051745W WO2022115973A1 WO 2022115973 A1 WO2022115973 A1 WO 2022115973A1 CA 2021051745 W CA2021051745 W CA 2021051745W WO 2022115973 A1 WO2022115973 A1 WO 2022115973A1
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cbda
thca
amine
amine salt
solvent
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Tony Durst
Jay VAN DER VLUGT
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Nectar Health Sciences Inc
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Nectar Health Sciences Inc
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    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01D—SEPARATION
    • B01D9/00—Crystallisation
    • B01D9/02—Crystallisation from solutions
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01D—SEPARATION
    • B01D11/00—Solvent extraction
    • B01D11/02—Solvent extraction of solids

Definitions

  • Various embodiments disclosed herein generally relate to methods for processing and separating mixtures of cannabinoid compounds extracted from crude complex extracts and mixtures. More specifically, this disclosure pertains to methods relating to solubilizing desolventized concentrated crude complex mixtures of metabolites and phytochemicals in selected solvents for separating, precipitating, and purifying cannabinoid compounds therefrom.
  • Cannabis is a genus of flowering plants in the Cannabaceae family.
  • Cannabis sp. are known to produce at least 113 distinct cannabinoids and over 50 terpenes that are concentrated in viscous resins produced in plant structures known as glandular trichomes. Trichomes are located at about the axial growing tips of Cannabis plants. Perhaps the most recognized cannabinoids are tetrahydrocannabinol (THC) and cannabidiol (CBD). It is well known that THC has significant but temporary psychoactive effects (i.e., hallucinogenic) on mammalian physiology and for this reason, various formats of Cannabis sp. plant materials and extracts are consumed for recreational use.
  • THC tetrahydrocannabinol
  • CBD cannabidiol
  • Cannabis terpenes are known to provide characteristic distinct aromas and flavors. It is also known that terpenes interact with cannabinoids to modulate the physiological effects of cannabinoids.
  • fiber-type cannabis commonly known as hemp
  • hemp has relatively high levels of CBD with very low levels or no levels of THC and consequently, is considered to have no or only minimal psychoactive and/or anxiogenic effects.
  • the term “hemp” derives its definition from legal and/or regulatory distinctions for fiber-type cannabis strains and cultivars that stably and reproducibly have less than 0.3% THC in the USA.
  • Cannabinoid compounds used for both recreational and medicinal purposes are almost exclusively crude extracts that have been solubilized and recovered from cannabis plant biomass with one of aqueous solvents, organic solvents, supercritical CO2, and the like as a first processing step.
  • the resulting crude extracts generally comprise complex mixtures of cannabinoids, terpenes, flavonoids, polyphenols, alkaloids, steroids, and other phytochemicals, which vary with the type of solvent selected for the extraction process. Numerous processes are known for use to refine crude complex extracts to separate out undesirable phytochemical components and to concentrate the cannabinoid and terpene components.
  • the chemical "fingerprint" of a particular botanical species can vary widely depending on the age of the plant, time of harvest, soil conditions, weather conditions, and a myriad of other factors. It is known that botanicals with very different phytochemical profiles will have different therapeutic effects, even if the botanicals are recovered from the same plant species. Standardization of botanical extraction processes facilitate the batch-to-batch reproducibility of a final product.
  • a standardized extract has a selected concentration of a marker compound that is known to a high degree of accuracy, and because both the amount of botanical material that is extracted and the amount of a carrier that may be added can be varied, it is possible to compensate for natural variability in the plant material. Also, if endogenous phytochemical active components of a standardized botanical extract are administered to patients in known quantities, then the treatments following prognosis of a disease can be monitored. Therefore, there is a need for standardized and reproducible extracts of botanicals, including extracts derived from cannabis.
  • the embodiments of the present disclosure generally relate to methods for separating, recovering, and purifying selected cannabinoid compounds from crude complex extracts and mixtures.
  • cannabis biomass feedstocks may be processed with one or more solvents selected from methanol, ethanol, propanol, isopropanol, butanol, propane, butane, ethyl acetate, acetone, dichloromethane, 1 ,4-dioxane, tetrahydrofuran, acetonitrile, toluene, methyl tert-butyl ether, supercritical CO2, subcritical CO2, hot water, supercritical H2O, subcritical H2O to recover therefrom crude extract suspensions in the selected solvent.
  • the crude extract suspension may be desolventized by removal of the extractant solvent to thereby produce a concentrated crude extract in the form of an oil or a resin or a solid.
  • fermentation broths wherein genetically modified microorganisms have been cultured to produce target cannabinoid phytochemicals may be processed to separate and recover therefrom complex mixtures of fermentation metabolites and phytochemicals.
  • the processing may include separating the cultured microorganisms from the fermentation broths prior to separation and recovery of complex mixtures of fermentation metabolites and phytochemicals.
  • the processing may include steps to recover metabolites and phytochemicals from the cultured microorganisms.
  • the complex mixtures may be processed by one or more of dewatering steps, desolventizing steps, drying steps, filtration steps including microfiltration steps, extraction with supercritical CO2 steps, and the like known to those skilled in this art, to produce a concentrated crude complex mixture in an oil form or a solid form or a dry form.
  • the concentrated crude extract is resolubilized in a solvent selected from methanol, ethanol, denatured ethanol, propanol, isopropanol, butanol, dichloromethane, ethyl acetate, C5-C7 alkanes and petroleum ethers, that is, the solvent selected for resolubilizing the concentrated crude complex extract is switched from the selected extractant solvent to thereby produce a solvent-switched crude complex extract solution.
  • a solvent selected from methanol, ethanol, denatured ethanol, propanol, isopropanol, butanol, dichloromethane, ethyl acetate, C5-C7 alkanes and petroleum ethers that is, the solvent selected for resolubilizing the concentrated crude complex extract is switched from the selected extractant solvent to thereby produce a solvent-switched crude complex extract solution.
  • solvent-switching refers to precipitation and recovery of cannabinoid carboxylic amine salts from solvent-solubilized concentrated crude complex extracts in solvents that are different from the extractant solvents that were used to extract and recover the concentrated crude complex extracts and mixtures.
  • Some embodiments pertain to methods for separating, recovering, and purifying one or more tetrahydrocannabinol phytochemicals (referred to herein as “THCs”) from crude complex extracts prepared from solvent-switched crude extract solutions according to the present disclosure.
  • THCs tetrahydrocannabinol phytochemicals
  • Some embodiments disclosed herein relate to methods for the use of selected amines to precipitate THCA-amine salts from solvent-switched crude complex extract solutions.
  • the precipitated THCA-amine salt may be washed one or more times with a selected alkane solvent and recrystallized to produce a purified THCA-amine salt.
  • a suitable amine for precipitating a THCA- amine salt from a solvent-switched crude complex extract may be an amino alcohol such as dimethylethanolamine (DMEA), 1 -piperidineethanol, and the like.
  • a suitable amine for precipitating a THCA- amine salt from a solvent-switched crude complex extract may a diamine such as 1 ,5-diazabicyclo[4.3.0]non-5-ene (DBN), and the like.
  • a purified THCA-amine salt produced with the methods disclosed herein may be decarboxylated to thereby produce a highly purified THC.
  • a purified THCA-amine salt produced with the methods disclosed herein may be acidified to thereby produce a highly purified THCA.
  • Some embodiments pertain to methods for forming, recovering, and purifying cannabidiolic acid-amine salts (CBDA-amine salts) from crude complex extracts prepared from solvent-switched crude extract solutions according to the present disclosure.
  • CBDA-amine salts cannabidiolic acid-amine salts
  • Some embodiments disclosed herein relate to methods for the use of selected amines to produce purified CBDA-amine salts.
  • a selected amine may be added to a solvent-switched crude complex extract to precipitate therefrom a CBDA-amine salt.
  • the precipitated CBDA-amine salt may be washed one or more times with a selected alkane solvent, and recrystallized to produce a purified CBDA-amine salt.
  • a suitable amine for precipitating a CBDA- amine salt from a solvent-switched crude complex extract may be selected from a tertiary amine such as triethylamine, tributylamine, N,N-diisopropylethylamime (Hunig’s base), and the like.
  • a suitable amine for precipitating a CBDA- amine salt from a solvent-switched crude complex extract may be selected from a diamine such 1 ,5-diazabicyclo[4.3.0]non-5-ene (DBN), 1 ,4- diazabicyclo[2.2.2]octane (DABCO), and the like.
  • a diamine such 1 ,5-diazabicyclo[4.3.0]non-5-ene (DBN), 1 ,4- diazabicyclo[2.2.2]octane (DABCO), and the like.
  • a purified CBDA-amine salt produced with the methods disclosed herein may be decarboxylated to thereby produce a highly purified CBD.
  • a purified CBDA-amine salt produced with the methods disclosed herein may be acidified to thereby produce a highly purified CBDA.
  • FIG. 1 A is a chart showing a linear calibration curve for cannabidivarin (CBDV);
  • FIG. 1 B is a chart showing a linear calibration curve for tetrahydrocannbidivarin (THCV);
  • FIG. 1C is a chart showing a linear calibration curve for cannabidiol (CBD).
  • FIG. 2A is a chart showing a linear calibration curve for cannabigerol (CBD).
  • FIG. 2B is a chart showing a linear calibration curve for cannabidiolic acid (CBDA);
  • FIG. 2C is a chart showing a linear calibration curve for cannabigerolic acid (CBDA);
  • FIG. 3A is a chart showing a linear calibration curve for cannabinol (CBN);
  • FIG. 3B is a chart showing a linear calibration curve for A 9 - tetrahydrocannabinol (A 9 -THC);
  • FIG. 3C is a chart showing a linear calibration curve for A 8 - tetrahydrocannabinol (A 8 -THC);
  • FIG. 4A is a chart showing a linear calibration curve for cannabichromene (CBC).
  • FIG. 4B is a chart showing a linear calibration curve for (-)-Trans-A 9 - tetrahydrocannabinolic acid (THCA);
  • FIG. 5 is an HPLC chromatogram showing separation of a standardized reference mixture of the eleven cannabinoid phytochemicals shown in FIGs. 1A- 4B;
  • FIG. 6 is an HPLC chromatogram showing a washed crude THCA-DMEA salt precipitated and recovered from the solvent-solubilized crude complex extract from Example 2;
  • FIG. 7 is an HPLC chromatogram showing a purified THCA-DMEA salt after recrystallization of the crude THCA-DMEA salt shown in FIG. 6 from Example 2;
  • FIG. 8 is an HPLC chromatogram showing a highly purified THCA after washing and removal of DMEA from purified THCA-DMEA salt shown in FIG. 7 in Example 2;
  • FIG. 9 is an HPLC chromatogram showing a washed crude THCA-DMEA salt precipitated and recovered from the solvent-solubilized crude complex extract from Example 3;
  • FIG. 10 is an HPLC chromatogram showing a washed and dried primary and secondary THCA-amine salts recovered from washed crude THCA-DMEA salt shown in FIG. 9 from Example 3;
  • FIG. 11 is an HPLC chromatogram showing a washed and dried highly purified THCA-DMEA salt recovered from the primary and secondary THCA- amine salts shown in FIG. 10 from Example 3;
  • FIG. 12 is an HPLC chromatogram showing a highly purified THCA solid produced from the highly purified THCA-DMEA salt shown in FIG. 11 from Example 3;
  • FIG. 13 is an HPLC chromatogram showing the cannabinoid content of a butane-desolventized crude Cannabis sativa extract resolubilized in heptane, from Example 4;
  • FIG. 14 is an HPLC chromatogram showing the cannabinoid content of the standard-solubilized crude C. sativa extract prepared from the extract shown in FIG. 13 from Example 4;
  • FIG. 15 is an HPLC chromatogram showing a purified THCA-DMEA salt precipitated from the standard-solubilized crude C. sativa extract shown in FIG. 14 from Example 4;
  • FIG. 16 is an HPLC chromatogram showing a purified solid THCA prepared from the purified THCA-DMEA salt shown in FIG. 15 from Example 4;
  • FIG. 17 is an HPLC chromatogram showing the cannabinoid content of the standard-solubilized crude C. sativa crude complex extract dissolved in 1- butanol from Example 5;
  • FIG. 18 is an HPLC chromatogram showing a purified THCA-DMEA salt precipitated from the standard-solubilized crude C. sativa extract shown in FIG. 17 from Example 5;
  • FIG. 19 is an HPLC chromatogram showing a purified THCA-1- piperidineethanol salt precipitated from the standard-solubilized crude C. sativa extract shown in FIG. 19 from Example 5;
  • FIG. 20 is an HPLC chromatogram showing the cannabinoid content of the standard-solubilized crude C. sativa crude complex extract dissolved in ethyl acetate from Example 6;
  • FIG. 21 is an HPLC chromatogram showing a purified THCA-DMEA salt precipitated from the standard-solubilized crude C. sativa extract shown in FIG. 20 from Example 6;
  • FIG. 22 is an HPLC chromatogram showing a purified THCA-DBN salt precipitated from the standard-solubilized crude C. sativa extract shown in FIG. 20 from Example 6;
  • FIG. 23 is an HPLC chromatogram showing a washed crude CBDA-TEA salt precipitated and recovered from the solvent-solubilized crude complex extract from Example 7;
  • FIG. 24 is an HPLC chromatogram showing a purified CBDA-TEA salt produced from the crude CBDA-TEA salt shown in FIG. 23 from Example 7;
  • FIG. 25 is an HPLC chromatogram showing a decarboxylated CBD oil produced from the purified CBDA-TEA salt shown in FIG. 24 from Example 7;
  • FIG. 26 is an HPLC chromatogram showing a highly purified CBDA solid produced from the purified CBDA-TEA salt shown in FIG. 24 from Example 7;
  • FIG. 27 is an HPLC chromatogram showing the cannabinoid content of concentrated crude complex extract produced from dried hemp biomass extracted with 95% ethanol, and resolubilized and standardized in 1 -butanol, from Example 8;
  • FIG. 28 is an HPLC chromatogram showing a purified CBDA-TEA salt precipitated from the standard-solubilized crude hemp extract shown in FIG. 27 from Example 8;
  • FIG. 29 is an HPLC chromatogram showing a purified CBDA-Hunig’s base amine salt precipitated from the standard-solubilized crude hemp extract shown in FIG. 27 from Example 8;
  • FIG. 30 is an HPLC chromatogram showing a purified CBDA-DABCO salt precipitated from the standard-solubilized crude hemp extract shown in FIG. 27 from Example 8;
  • FIG. 31 is an HPLC chromatogram of an unfiltered HPLC-methanol control sample in Example 8, Example 9, Example 10, and Example 11 ;
  • FIG. 32 is an HPLC chromatogram of a filtered HPLC-methanol control sample showing an unknown anomaly associated with the filter labeled as “THC- A in Example 8, Example 9, Example 10, and Example 11 ;
  • FIG. 33 is an HPLC chromatogram showing the cannabinoid content of the standardized reconstituted purified CBDA dissolved in 1 -butanol, from Example 9;
  • FIG. 34 is an HPLC chromatogram showing a purified CBDA-triethylamine salt precipitated from the CBDA-1 -butanol stock solution shown in FIG.33 from Example 9
  • FIG. 35 is an HPLC chromatogram showing a purified CBDA-Hunig’s base amine salt precipitated from the CBDA-1 -butanol stock solution shown in FIG. 33 from Example 9;
  • FIG. 36 is an HPLC chromatogram showing a purified CBDA-tributylamine salt precipitated from the CBDA-1 -butanol stock solution shown in FIG. 33 from Example 9;
  • FIG. 37 is an HPLC chromatogram showing a purified CBDA-DBN salt precipitated from the CBDA-1 -butanol stock solution shown in FIG. 33 from Example 9;
  • FIG. 38 is an HPLC chromatogram showing a purified CBDA-DABCO salt precipitated from the CBDA-1 -butanol stock solution shown in FIG. 33 from Example 9;
  • FIG. 39 is an HPLC chromatogram showing a purified CBDA-1- piperidineethanol amine salt precipitated from the CBDA-1 -butanol stock solution shown in FIG. 33 from Example 9;
  • FIG. 40 is an HPLC chromatogram showing the cannabinoid content of concentrated crude complex extract produced from dried hemp biomass extracted with 95% ethanol, and resolubilized and standardized in ethyl acetate, from Example 10;
  • FIG. 41 is an HPLC chromatogram showing a purified CBDA-triethylamine salt precipitated from the CBDA-ethyl acetate stock solution shown in FIG.40 from Example 10;
  • FIG. 42 is an HPLC chromatogram showing a purified CBDA-DMEA amine salt precipitated from the CBDA-ethyl acetate stock solution shown in FIG.40 from Example 10;
  • FIG. 43 is an HPLC chromatogram showing a purified CBDA-Hunig’s base amine salt precipitated from the CBDA-ethyl acetate stock solution shown in FIG. 40 from Example 10;
  • FIG. 44 is an HPLC chromatogram showing the cannabinoid content of concentrated crude complex extract produced from dried hemp biomass extracted with denatured ethanol, and resolubilized and standardized in denatured ethanol, from Example 11 ;
  • FIG. 45 is an HPLC chromatogram showing a purified CBDA-triethylamine salt precipitated from the CBDA-denatured ethanol stock solution shown in FIG.44 from Example 11 ;
  • FIG. 46 is an HPLC chromatogram showing a purified CBDA-Hunig’s base amine salt precipitated from the CBDA-denatured ethanol stock solution shown in FIG. 44 from Example 11 .
  • any atom shown in a drawing with unsatisfied valences is assumed to be attached to enough hydrogen atoms to satisfy the valences.
  • chemical bonds depicted with one solid line parallel to one dashed line encompass both single and double (e.g., aromatic) bonds, if valences permit.
  • the singular forms “a”, “an”, and “the,” may also refer to plural articles, i.e., “one or more”, “at least one”, “and/or”, are open-ended expressions that are both conjunctive and disjunctive in operation.
  • a cannabinoid includes “one or more cannabinoids”.
  • each of the expressions “at least one of A, B, and C", “at least one of A, B, or C", “one or more of A, B, and C", “one or more of A, B, or C” and “A, B, and/or C” means A alone, B alone, C alone, A and B together, A and C together, B and C together, or A, B and C together.
  • an entity refers to one or more of that entity. As such, the terms “a”, “an”, “one or more”, and “at least one” can be used interchangeably herein.
  • the terms “cannabis” and “cannabis biomass” encompass whole Cannabis sativa plants and also parts thereof which contain cannabinoids and cannabis phytochemicals, such as the aerial parts of the plants or isolated leaves and/or flowering heads and/or seeds.
  • the term also encompasses freshly harvested cannabis plant material and also plant material, cannabis plant material that was dried after harvesting.
  • Dried cannabis plant material may be in a loose form or alternatively, may be baled into square bales or rectangular bales or round bales or alternatively, may be compressed into cubes or pellets or cubes.
  • Dried cannabis plant material may be separated into two or more components wherein one component comprises the cannabis stalks and stems, and a second component comprises the leaves, trichomes, and flowers.
  • the second component may be further separated into leaves and trichome/flower components and the trichome/flower components may be separated into trichome and flower components.
  • kief refers to a powdery resin that is produced by and may be collected from the trichomes of cannabis plants.
  • cannabinoid encompasses cannabidiol (CBD), cannabidiolic acid (CBDA), cannabinol (CBN), cannabigerol (CBD), cannabigerolic acid (CBDA), cannabichromene (CBC), cannabichromenic (CBCA), cannabicyclol (CBL), cannabivarin (CBV), cannabidivarin (CBDV), cannabidivarinic (CBDVA), cannabichromevarin (CBCV), cannabigerovarin (CBDV), cannabigerol monomethyl ether (CBDM), cannabielsoin (CBE), cannabicitran (CBT), among others.
  • CBD cannabidiol
  • CBD cannabidiolic acid
  • CBD cannabinol
  • CBD cannabigerol
  • CBDA cannabigerolic acid
  • CBC cannabichromene
  • CBCA cannabichromen
  • tetrahydrocannabinol encompasses (-)-trans-A9-tetrahydrocannabinol (A9-THC), A8- tetrahydrocannabinol (A8-THC), iso-tetrahydrocannabinol, tetrahydrocannabinolic acid (THCA), tetrahydrocannabivarin (THCV), tetrahydrocannabivarinic acid (THCVA), among others.
  • THCA tetrahydrocannabinolic acid
  • THCV tetrahydrocannabivarin
  • THCVA tetrahydrocannabivarinic acid
  • tetrahydrocannabinol may also be substituted for herein by the acronym “THC”.
  • cinnamonbis phytochemicals refers to biologically active compounds produced by Cannabis sativa plants, and in particular, to mixtures of terpenes, terpenoids, flavonoids, alkaloids, lignans, omega fatty acids, pigments, and the like, that may be extracted and separated from cannabis biomass by solvent extraction.
  • phytochemical refers to a single biologically active compound that has been separated from a mixture of phytochemicals.
  • fermentation broths wherein genetically modified microorganisms have been cultured to produce target cannabinoid phytochemicals may be processed to separate and recover therefrom complex mixtures of fermentation metabolites and phytochemicals.
  • the processing may include separating the cultured microorganisms from the fermentation broths prior to separation and recovery of complex mixtures of fermentation metabolites and phytochemicals.
  • the processing may include steps to recover metabolites and phytochemicals from the cultured microorganisms.
  • the complex mixtures may be processed by one or more of dewatering steps, desolventizing steps, drying steps, filtration steps including microfiltration steps, extraction with supercritical CO2 steps, and the like known to those skilled in this art, to produce a concentrated crude complex mixture in an oil form or a solid form or a dry form.
  • Suitable methods for preparing complex mixtures of fermentation metabolites and phytochemicals from fermentation broths and from cultured microorganism may be found in International Patent Application Publication No. WO 2020/160284A1 , International Patent Application Publication No. WO 2020/176998CA, IUS Patent Application Publication No. 2021/0189444A1 , Chapter 10 “The recovery and purification of fermentation products” in the third edition of “Principles of Fermentation Technology” (Stanbury et al., 2016).
  • solvent as used herein, is used herein to denote a liquid or gas capable of dissolving a solid or another liquid or gas.
  • solvents include alcohols such as methanol, ethanol, propanol, isopropanol, butanol, propane, butane, low boiling point (b.p.) petroleum ethers, ethyl acetate, acetone (also known as propanone), dichloromethane, 1 ,4-dioxane, tetrahydrofuran, acetonitrile, toluene, methyl tert-butyl ether, supercritical carbon dioxide (CO2), subcritical CO2, and the like.
  • alcohols such as methanol, ethanol, propanol, isopropanol, butanol, propane, butane, low boiling point (b.p.) petroleum ethers, ethyl acetate, acetone (also known as propanone), dichloromethane, 1 ,4-
  • solvent switching refers to a process for extracting, separating, recovering, and purifying selected cannabinoids from cannabis biomass wherein the first step is to process a cannabis biomass with a first solvent selected from methanol, ethanol, propanol, isopropanol, butanol, propane, butane, ethyl acetate, acetone, dichloromethane, 1 ,4-dioxane, tetrahydrofuran, acetonitrile, toluene, methyl tert-butyl ether, supercritical carbon dioxide (CO2), subcritical CO2, and the like, to produce a crude complex extract.
  • a first solvent selected from methanol, ethanol, propanol, isopropanol, butanol, propane, butane, ethyl acetate, acetone, dichloromethane, 1 ,4-dioxane, tetrahydrofuran, acetonitrile, tol
  • the next step is to desolventize the crude complex extract to recover the extractant solvent therefrom to produce a concentrated cannabis extract in the form of an oil or a resin or a solid.
  • the next step is to resolubilize the concentrated cannabis extract in a second solvent (that is, switching the processing solvents) selected from one of methanol, ethanol, denatured ethanol, propanol, isopropanol, butanol, dichloromethane, ethyl acetate, C5-C7 low- boiling hydrocarbon solvents including alkanes and petroleum ethers to thereby produce a solvent-switched crude complex extract.
  • a second solvent that is, switching the processing solvents
  • solvent switching also refers to a process for extracting, separating, recovering, and purifying selected cannabinoids from fermentation systems wherein selected genetically modified microorganisms were cultured therein to produce a selected targeted cannabinoid, wherein the first step is to process the spent fermentation broths with a first solvent selected from methanol, ethanol, propanol, isopropanol, butanol, propane, butane, ethyl acetate, acetone, dichloromethane, 1 ,4-dioxane, tetrahydrofuran, acetonitrile, toluene, methyl tert-butyl ether, supercritical carbon dioxide (CO2), subcritical CO2, and the like, to produce a first crude complex mixture.
  • a first solvent selected from methanol, ethanol, propanol, isopropanol, butanol, propane, butane, ethyl acetate, acetone, dichloromethane
  • the selected genetically modified microorganisms may be processed with the selected first solvent to produce a second crude complex mixture.
  • the first and second crude complex mixtures may be combined to recover the extractant solvent therefrom to produce a concentrated crude mixture in the form of an oil or a resin or a solid.
  • the extractant solvent may be recovered separately from each of the first crude complex mixture and the second crude complex mixture to produce therefrom concentrated crude complex mixtures.
  • the next step is to resolubilize the concentrated complex mixtures in a second solvent (that is, switching the processing solvents) selected from one of methanol, ethanol, denatured ethanol, propanol, isopropanol, butanol, dichloromethane, ethyl acetate, C5-C7 low-boiling hydrocarbon solvents including alkanes and petroleum ethers to thereby produce solvent-switched crude complex mixtures containing therein the selected targeted cannabinoid.
  • a second solvent that is, switching the processing solvents
  • the term “antisolvent” refers to an organic solvent that may be used to precipitate a target compound or molecule from another solvent in which the target compound or molecule is completely dissolved whereby, as the antisolvent is added to the solvent containing the dissolved target compound or molecule, the precipitation process is initiated by nucleation of the target compound or molecule followed by the formation of solid particles.
  • an alcohol was a solvent selected for dissolution of a target compound or molecule
  • water may be a suitable antisolvent to precipitate the target compound or molecule.
  • crude complex extract refers to a complex mixture of cannabis phytochemicals that were extracted, then separated and recovered from cannabis biomass by processing of the biomass with a selected solvent.
  • the term “crude complex extract” also refers to concentrated crude complex mixtures recovered and prepared from spent fermentation broths wherein genetically modified microorganisms were cultured to produce selected targeted cannabinoid compounds and from the genetically modified microorganisms.
  • the crude complex extracts may be in a resin form or a gum form or an oil form.
  • the term “crude precipitate” as used herein means the solids and/or oils produced by a chemical reaction between a selected organic base with a mixture of cannabinoid carboxylic acids present in a crude complex extract.
  • the “crude precipitate” may also be referred to herein as a “crude isolate” or a “carboxylic acid salt” or a “precipitated cannabinoid”.
  • purified precipitate means the solids and/or oils remaining after the crude precipitate is washed with a selected solvent such as, for example, with ethyl acetate at 40 °C.
  • a purified precipitate may also be produced via a recrystallization process wherein the crude precipitate is dissolved in a heated solvent and then cooled to an appropriate temperature to induce crystallization.
  • the crude precipitate may be dissolved in a solvent which readily dissolves both the desired purified precipitate and the impurities present in the crude precipitate, followed by addition of an antisolvent in which the desired precipitate is insoluble and the impurities remain in solution. Subsequent filtration yields the purified precipitate.
  • the “purified precipitate” may also be referred as a “purified isolate” or a “purified cannabinoid precipitate” or a “purified cannabinoid carboxylic acid”.
  • a “standardized solvent-solubilized crude extract” refers to a crude extract that has been adjusted by the addition or removal of a solvent to adjust the concentrations therein of one or more bioactive markers, such as THCA or CBDA, to a selected target range in comparison to the concentrations of the one or more bioactive markers in a reference solution, using analytical methods known to those skilled in these arts.
  • suitable analytical methods include HPLC methods and the like.
  • Some embodiments disclosed herein relate to methods of separating and recovering CBDA or THCA from solubilized crude complex extracts comprising cannabinoids and other phytochemicals extracted and recovered from cannabis biomass feedstocks or from fermentation systems wherein selected genetically modified microorganisms were cultured to produce selected target cannabinoid compounds.
  • the methods for specifically separating and recovering CBDA or THCA from solubilized crude complex extracts pertain to the use of one or more selected amines to selectively react with CBDA or THCA thereby forming CBDA-amine salts or THCA- amine salts that precipitate out of the crude complex solutions.
  • the methods disclosed herein include steps for separating and recovering precipitated CBDA-amine salts or THCA-amine salts from crude complex extract solutions, for washing recovered CBDA-amine salts or THCA-amine salts to separate and remove therefrom other cannabinoids and cannabis phytochemicals that may have been recovered with the precipitated CBDA- amine salts or THCA-amine salts, for further purifying and recrystallization of the washed THCA-amine salts or CBDA-amine salts, for the preparation of purified crystalline CBDA or THCA, and for decarboxylating the purified THCA-amine salts or CBDA-amine salts to produce purified THC or CBD therefrom.
  • solvent selected from methanol, ethanol, denatured ethanol, propanol, isopropanol, butanol, dichloromethane, ethyl acetate, C5-C7 low-boiling hydrocarbon solvents including alkanes and petroleum ethers, certain amines preferentially precipitated CBDA salts while certain other amines preferentially precipitated THCA salts.
  • the amine-precipitated CBDA salts also referred to herein as CBDA-amine salts, have low or very low solubility in a number of organic solvents at room temperature and therefore, may be washed with those organic solvents to produce highly purified CBD-amine salts.
  • the amine- precipitated THCA salts also referred to herein as THCA-amine salts, have low or no solubility in a number of organic solvents at room temperature and therefore, may be washed with those organic solvents to produce highly purified THC-amine salts.
  • the structure of the precipitated THCA- amine salt produced in the above reaction can be verified by 1 H NMR spectroscopy which shows the expected 6: 1 :1 ratio of the hydrogen nuclei due to the dimethylamino group [2.6ppm, s, 6H] present in the basic component, the remaining single hydrogen ion on the aromatic ring [6.2 ppm, s, 1 H] and the alkene H [6.8ppm, broad s, 1 H] due to the THC moiety.
  • the precipitated THCA-amine salt may be recrystallized and purified by (i) first slurrying and at least partially dissolving the salt in a selected volume of a solvent such as, for example, ethyl acetate or heptane or a mixture thereof, and then (ii) slowly adding a selected suitable antisolvent such as hexane or heptane or pentane or petroleum ether until recrystallization of the THCA into a purified salt form commences.
  • a solvent such as, for example, ethyl acetate or heptane or a mixture thereof
  • the precipitated THCA-amine salt can be dissolved in a minimum of a suitable organic solvent by heating, and the cooling to thereby cause recrystallization to occur.
  • the recrystallized THCA salt may be dissolved in a suitable polar aprotic solvent, for example ethyl acetate, to which is added a 0.1 M-HCI solution to cause partitioning of the mixture into an aqueous layer and an organic solvent layer.
  • a suitable polar aprotic solvent for example ethyl acetate
  • Sufficient 0.1 M-HCI solution is added until the aqueous layer turns litmus paper a red color.
  • the two resulting layers may be separated using separation processes known to those skilled in this art.
  • HPLC analysis of the partitioned ethyl acetate layer will show the presence of very high-purity A 9 -THCA (in reference to a A 9 -THCA standard).
  • Ethyl acetate may be removed from the partitioned organic layer thereby producing very high-purity A 9 -THCA initially as an oil that solidifies into a powder form.
  • crystallization is an important step for separation and purification of THCA. Crystallization involves two key steps: (i) formation of solid particles from liquid solution (nucleation) and (ii) growth due to the deposition of additional substances on existing particles.
  • the thermodynamic driving force behind both steps is the difference in chemical potential between the solution phase that is liquid phase and the crystal phase that is solid phase.
  • the difference can be represented by supersaturation, which is defined as the difference between the actual concentration of the crystallizing substance in the solution and its saturation concentration.
  • crystallization of cannabinoids in a solvent mixture is a function of temperature and time.
  • a way to induce crystallization is to lower the temperature below the saturation point of a particular constituent that can then precipitate out of the solution as a solid. Crystallization may also be induced by providing seed crystals to the mixture and/or by scratching an inner surface of the vessel wherein the mixture is contained.
  • Suitable amine bases in addition to DMEA, that may be used in the methods disclosed herein to precipitate THCA-amine salts from solvent-switched crude complex extracts, include 1 ,5-diazabicyclo(4.3.0)non-5-ene (DBN), 1 -piperidineethanol, and the like.
  • An embodiment of the present disclosure pertains to an example method for separating out, recovering, and purifying THCA in the form of a THCA-amine salt, from a crude extract comprising a mixture of cannabinoids and cannabis phytochemicals recovered from processing cannabis biomass.
  • the example method comprises the steps of: 1 . providing a desolventized concentrated crude complex extract in resin form or an oil form or a solid form;
  • a suitable target range for adjusting the THCA content to in step 2 may be from about 20 mg/mL to about 445 mg/mL.
  • a particularly suitable target range may be from about 27 mg/mL to about 200 mg/rnL.
  • a preferred target range may be from about 31 mg/rnL to about 153 mg/rnL.
  • the standardized solvent-solubilized crude extract may be spiked with a selected volume of a denatured ethanol prior to step 3 of adding and mixing the selected amine thereinto.
  • a suitable volume of denatured ethanol may be selected from a range of about 2% to about 20% by volume of the standardized solvent-solubilized crude extract.
  • the standardized solvent-solubilized crude extract may be spiked with a selected volume of acetone prior to adding and mixing the selected amine thereinto.
  • a suitable volume of acetone may be selected from a range of about 4% to about 20% by volume of the standardized solvent-solubilized crude extract.
  • a suitable second organic solvent for washing the recovered crude THCA-amine salt in step 5 may be a C5-C7 alkane or a low b.p. petroleum ether.
  • Particularly suitable alkanes may be heptane, hexane, and pentane.
  • a suitable third organic solvent for resolubilizing the washed crude THCA-amine salt in step 6 may be one of ethyl acetate, 85%-99% ethanol, denatured ethanol, methanol, isopropanol, dichloromethane, toluene, MTBE, THF, and the like.
  • a particularly suitable solvent for resolubilizing the washed CBDA-amine salt in step 6, may be ethyl acetate heated to about 76 °C.
  • a suitable antisolvent for recrystallizing the solubilized THC-amine salt in step 7 may be an alkane such as one of heptane, hexane, pentane, and the like.
  • a suitable antisolvent may be water.
  • Another embodiment of the present disclosure pertains to an example method for separating out, recovering, and purifying a THCA from a purified THCA-amine salt produced by following the method steps 1 to 8 previously disclosed herein, additionally comprise the steps of: 9. re-solubilizing the purified THCA-amine salt in a selected fourth organic solvent, then
  • a suitable fourth organic solvent for use in step 9 may be one of ethyl acetate, heptane, hexane, pentane, low boiling point petroleum ethers, butanol, and dichloromethane.
  • a suitable mineral acid solution for use in step 10 may be one of 5% HCI, 5% H2SO4, and the like.
  • Another embodiment of the present disclosure pertains to an example method for separating out, recovering, and purifying THC from a THCA-amine salt, produced by following method steps 1 to 8 previously disclosed herein, additionally comprising the steps of:
  • the recrystallized purified THCA-amine salt may be decarboxylated in step 13, by adding the THCA-amine salt into a sodium carbonate (Na2COs) solution, then heating the mixture under constant mixing at a temperature selected from a range of about 98-102 °C to reflux for a period of time selected from a range of about 2 hr to about 18 hr, thereby producing a biphasic solution of THC oil and separated amine organic phase, and an aqueous phase containing the Na2COs solution.
  • a suitable concentration of Na2COs solution to use for this step is from a range of about 1% to about 15% (w/v).
  • a particularly suitable concentration of Na2COs solution is from a range of about 2.5% to about 10% (w/v), for example, about 5% (w/v).
  • a particularly suitable temperature for this decarboxylation step is about 100 °C.
  • a particularly suitable time duration for this decarboxylation step is about 4 hr.
  • a suitable fifth organic solvent for use in step 14 may be one of dichloromethane or C5 to C7 alkane and the like.
  • a suitable mineral acid solution for use in step 15 may a 5% HCI solution, a 5% H2SO4 solution, and the like.
  • THCA-amine salts that have been precipitated and recovered from solvent-solubilized crude complex extracts with an amine selected from one of DMEA, DBN, 1- piperidineethanol, and the like.
  • An example method for producing purified THCA-amine salts comprises the steps of:
  • DMEA may be added to and commingled with a solvent-solubilized crude complex extract to precipitate therefrom a THCA-amine salt having a chemical structure shown in (1 ):
  • the precipitated THCA-DMEA salt may be washed with a selected organic solvent to thereby produce a purified THCA-DMEA salt.
  • DBN may be added to and commingled with a solvent-solubilized crude C. sativa extract to precipitate therefrom a THCA-amine salt having a chemical structure shown in (2):
  • the precipitated THCA-DBN salt may be washed with a selected organic solvent to thereby produce a purified THCA-DBN salt.
  • 1 -piperidineethanol may be added to and commingled with a solvent-solubilized crude C. sativa extract at a temperature at or below 0 °C to precipitate therefrom a THCA-amine salt having a chemical structure shown in (3):
  • the precipitated THCA-1-piperidineethanol salt may be washed with a selected organic solvent to thereby produce a purified THCA-1 -piperidineethanol salt.
  • Addition to a resolubilized solvent-switched crude extract of an amine selected from triethylamine, N,N-diisopropylethylamine (Hunig's base), 1 ,5- diazabicyclo[4.3.0]non-5-ene (DBN), dimethylethanolamine (DMEA), 1- piperidineethanol, and the like may precipitate CBDA-amine salts from the resolubilized solvent-switched crude extract.
  • Addition to a resolubilized solvent- switched crude extract at a temperature of 0 °C or less, of an amine selected from 1 ,4-diazabicyclo[2.2. 2]octane (DABCO), tributylamine, and the like may precipitate CBDA-amine salts from the resolubilized solvent-switched crude extract.
  • CBDA- amine salts formed as an oil and/or precipitated as a solid salt by a selected amine as disclosed herein may be washed with a selected solvent to remove other cannabinoids and/or cannabis phytochemicals that may have remained associated with the recovered precipitated CBDA-amine salts.
  • Suitable solvents for the washing step may include C5-C7 alkanes, ethyl acetate, isopropanol, denatured ethanol, 95% ethanol, and 100% ethanol.
  • washed CBDA-amine salts may be further purified by addition and mixing into a heated mixture of a polar solvent and non-polar solvent to form a solution, and then, may be recrystallized back into a purified CBDA-amine salt by cooling or by the addition of an antisolvent.
  • a suitable polar solvent may be one of ethyl acetate, 95% ethanol, methanol, isopropanol, dichloromethane, toluene, methyl-tert-butyl ether (MTBE), tetrahydrofuran (THF), and the like.
  • a particularly suitable polar solvent/non- polar solvent is a mixture of ethyl acetate with heptane.
  • Suitable antisolvents for use with such solvents include C5-C7 alkanes and low boiling point (b.p.) petroleum ethers.
  • washed CBDA-amine salts may be solubilized in an alcohol such as denatured ethanol or methanol, and then, may be recrystallized back into a purified CBDA-amine salt by cooling or by the addition of water as the antisolvent.
  • crystallization is an important step for separation and purification of CBDA. Crystallization involves two key steps: (i) formation of solid particles from liquid solution (nucleation) and (ii) growth due to the deposition of additional substances on existing particles.
  • the thermodynamic driving force behind both steps is the difference in chemical potential between solution phase that is liquid phase and crystal phase that is solid phase.
  • the difference can be represented by supersaturation, which is defined as the difference between the actual concentration of the crystallizing substance in the solution and its saturation concentration.
  • crystallization of cannabinoids in a solvent mixture is a function of temperature and time.
  • a way to induce crystallization is to lower the temperature below the saturation point of a particular constituent that can then precipitate out of the solution as a solid. Crystallization may also be induced by providing seed crystals to the mixture and/or by scratching an inner surface of the vessel wherein the mixture is contained.
  • a suitable ratio for the polar solvent/non- polar solvent mixture may be selected from a range of about 5: 1 to about 20: 1 .
  • a particularly suitable polar solvent/non-polar solvent ratio may be about 10: 1 , for example 10 parts ethyl acetate and 1 part heptane.
  • the CBDA-amine salts/polar solvent/non-polar solvent slurry is then cooled to about 30 °C, and then may be placed into a 4 °C environment for a period of time selected from about 30 min to about 12 h during which time, purified CBDA-amine salt will recrystallize out of the polar solvent/non-polar solvent mixture.
  • the recrystallized purified CBDA-amine salt may then be separated from the polar solvent/non-polar solvent mixture, for example, by filtration or centrifugation.
  • purified CBDA-amine salts produced by the methods disclosed herein may be decarboxylated and then separated by acidification to thereby produce a purified CBD.
  • the CBDA-amine salts produced by the methods disclosed herein may be acidified to separate the amines therefrom to produce highly purified CBDA.
  • a concentrated crude complex extract in the form of a resin or an oil or a solid may be analyzed prior to dilution in a selected solvent, to determine its cannabinoid composition and to determine the content of CBDA therein on a mass basis. Then, the crude extract may be diluted with a solvent selected from one of methanol, ethanol, denatured ethanol, propanol, isopropanol, butanol, dichloromethane, ethyl acetate, 1 -butanol, ethyl acetate, and a C5-C7 alkane.
  • a particularly suitable solvent may be denatured ethanol, 1-butanol, ethyl acetate, dichloromenthane, and C5-C7 alkanes such as heptane, hexane, and the like.”
  • a suitable target range may be from about 20 mg/mL to about 445 mg/rnL. Particularly suitable target ranges may be from about 27 mg/mL to about 200 mg/mL. Preferred target ranges may be from about 31 mg/mL to about 153 mg/mL.
  • An embodiment of the present disclosure pertains to an example method for separating out, recovering, and purifying CBDA in the form of a CBDA-amine salt, that has been precipitated and recovered from a crude complex extract comprising a mixture of metabolites, cannabinoids, and cannabis phytochemicals recovered from cannabis biomass, or from a complex mixture of metabolites, cannabinoids, and cannabis phytochemicals recovered from cultured microbial fermentation systems, with a suitable selected amine.
  • the example method comprises the steps of:
  • a first solvent selected from one of from one of methanol, ethanol, denatured ethanol, propanol, isopropanol, butanol, dichloromethane, ethyl acetate, 1-butanol, ethyl acetate, and a C5-C7 alkane with the crude complex extract to reduce the CBDA content therein to a level within a selected target range in reference to a CBDA standard, thereby producing a standardized solvent-solubilized crude complex extract;
  • a suitable target range for adjusting the CBDA content in step 2 may be from about 20 mg/rnL to about 445 mg/rnL
  • a particularly suitable target range may be from about 27 mg/rnL to about 200 mg/rnL.
  • a preferred target range may be from about 31 mg/rnL to about 153 mg/rnL.
  • the standardized solvent-solubilized crude extract may be spiked with a selected volume of a denatured ethanol prior to step 3 of adding and mixing the selected amine thereinto.
  • a suitable volume of denatured ethanol may be selected from a range of about 2% to about 20% by volume of the standardized solvent-solubilized crude extract.
  • the standardized solvent-solubilized crude extract may be spiked with a selected volume of acetone prior to adding and mixing the selected amine thereinto.
  • a suitable volume of acetone may be selected from a range of about 4% to about 20% by volume of the standardized solvent-solubilized crude extract.
  • a suitable second organic solvent for washing the recovered crude CBDA-amine salt in step 5 may be a C5-C7 alkane or a low b.p. petroleum ether.
  • Particularly suitable alkanes may be heptane, hexane, and pentane.
  • a suitable third organic solvent for resolubilizing the washed crude CBDA-amine salt in step 6 may be one of ethyl acetate, 85%-99% ethanol, denatured ethanol, methanol, isopropanol, dichloromethane, toluene, MTBE, THF, and the like.
  • a particularly suitable solvent for resolubilizing the washed CBDA-amine salt in step 6, may be ethyl acetate heated to about 76 °C.
  • a suitable antisolvent for recrystallizing the solubilized CBDA-amine salt in step 7 may be an alkane such as one of heptane, hexane, pentane, and the like.
  • a suitable antisolvent may be water.
  • Another embodiment of the present disclosure pertains to an example method for separating out, recovering, and purifying a CBDA from a purified CBDA-amine salt produced by following the method steps 1 to 8 previously disclosed herein, additionally comprise the steps of:
  • a suitable fourth organic solvent for use in step 9 may be one of ethyl acetate, hexane, heptane, pentane, low boiling point petroleum ethers, butanol, and dichloromethane.
  • a suitable mineral acid solution for use in step 10 may be one of 5% HCI, 5% H2SO4, and the like.
  • Another embodiment of the present disclosure pertains to an example method for separating out, recovering, and purifying CBD from a CBDA-amine salt, produced by following method steps 1 to 8 previously disclosed herein, additionally comprising the steps of:
  • the recrystallized purified CBDA-amine salt may be decarboxylated in step 13, by adding the CBDA-amine salt into a sodium carbonate (Na2COs) solution, then heating the mixture under constant mixing at a temperature selected from a range of about 98-102 °C to reflux for a period of time selected from a range of about 2 hr to about 18 hr, thereby producing a biphasic solution of CBD oil and separated amine organic phase, and an aqueous phase containing the Na2COs solution.
  • a suitable concentration of Na2CC>3 solution to use for this step is from a range of about 1% to about 15% (w/v).
  • a particularly suitable concentration of Na2COs solution is from a range of about 2.5% to about 10% (w/v), for example, about 5% (w/v).
  • a particularly suitable temperature for this decarboxylation step is about 100 °C.
  • a particularly suitable time duration for this decarboxylation step is about 4 hr.
  • a suitable fifth organic solvent for use in step 14 may be one of dichloromethane or C5 to C7 alkane and the like.
  • a suitable mineral acid solution for use in step 15 may a 5% HCI solution, a 5% H2SO4 solution, and the like.
  • triethylamine may be added to and commingled with a resolubilized solvent-switched crude complex extract to precipitate therefrom a CBDA-amine salt having a chemical structure shown in (4):
  • the precipitated CBDA-triethylamine salt may be washed with a selected organic solvent, optionally recrystallized, and then dried to thereby produce a purified CBDA-triethylamine salt.
  • N,N-diisopropylethylamine may be added to and commingled with a solvent-solubilized crude complex extract to precipitate therefrom a CBDA-amine salt having a chemical structure shown in (10):
  • the precipitated CBDA-N,N-diisopropylethylamine salt may be washed with a selected organic solvent, optionally recrystallized, and then dried to thereby produce a purified CBDA-diisopropylethylamine salt.
  • N,N-dimethylethanolamine (DMEA) may be added to and commingled with a solvent-solubilized crude complex extract to precipitate therefrom a CBDA-amine salt having a chemical structure shown in (6):
  • the precipitated CBDA-dimethylethanolamine salt may be washed with a selected organic solvent, optionally recrystallized, and then dried to thereby produce a purified CBDA-dimethylethanolamine salt.
  • 1 ,5-diazabicyclo[4.3.0]non-5-ene may be added to and commingled with a solvent-solubilized crude complex extract to precipitate therefrom a CBDA-amine salt having a chemical structure shown in (7):
  • 1 -piperidineethanol may be added to and commingled with a solvent-solubilized crude complex extract to precipitate therefrom a CBDA-amine salt having a chemical structure shown in (8):
  • the precipitated CBDA-1 -piperidineethanol salt may be washed with a selected organic solvent, optionally recrystallized, and then dried to thereby produce a purified CBDA-1 -piperidineethanol salt.
  • 1 ,4-diazabicyclo[2.2.2]octane (DABCO) (diamine) may be added to and commingled with a solvent-solubilized crude complex extract to precipitate therefrom a CBDA-amine salt having a chemical structure shown in (9):
  • the precipitated CBDA-DABCO salt may be washed with a selected organic solvent, optionally recrystallized, and then dried to thereby produce a purified CBDA-DABCO salt.
  • tributylamine may be added to and commingled with a solvent-solubilized crude complex extract at temperatures below 0 °C to precipitate therefrom a CBDA-amine salt having a chemical structure shown in (10):
  • the precipitated CBDA-tributylamine salt may be washed with a selected organic solvent, optionally recrystallized, and then dried to thereby produce a purified CBDA-tributylamine salt.
  • CBD cannabidivarin
  • THCV cannabidiol
  • CBD cannabigerol
  • CBD cannabidiolic acid
  • CBDA cannabigerolic acid
  • CBD cannabinol
  • CBC cannabichromene
  • a 8 - tetrahydrocannabinolic acid A 8 -THCA.
  • FIG. 1B THCV
  • FIG. 1C CBD
  • FIG. 2A CBD
  • FIG. 2B CBD-A
  • FIG. 2C CBDA
  • FIG. 3A CBN, FIG. 3B, A 9 -THC
  • FIG. 3C A 8 -THC
  • FIG. 4A CBC
  • FIG. 4B THCA.
  • a mixture containing 22.73 pg/mL of each of the eleven above-noted cannabinoid phytochemical compounds was prepared and then analyzed with the Agilent 1220 Infinity II LC System. The HPLC analysis of the mixture is shown in FIG. 5 and summarized below in Table 1.
  • CBD-A 5.830 12.32 125.316 25.0633
  • EXAMPLE 2 Cannabis sativa biomass was extracted with butane to produce a crude extract comprising a complex mixture of plant metabolites and phytochemicals including cannabinoid compounds. After separation from the extracted biomass, the crude complex extract was desolventized to produce a concentrated crude complex extract in the form of a resin. 103.62 g of the desolventized concentrated complex extract was dissolved in 1.38 L of n-heptane to produce a normalized solvent-solubilized extract solution containing approximately 64mg/mL of THCA.
  • a sample of the crude THCA-DMEA salt was solubilized in methanol, diluted 250X, and analyzed by HPLC (FIG. 6).
  • the washed and dried THCA amine salt was then recrystallized by dissolving 53.90 grams of THCA- DMEA salt in 3:1 volume/mass ratio ethyl acetate (162mL) under refluxing conditions and then cooled slowly to 36 °C at which temperature the rate of precipitation of solid THCA amine salt increased rapidly and a 1 :1 volume ratio of warm heptane(162 mL, 28 °C) was added to the recrystallization mixture. The mixture was further cooled to 4 °C and held at this temperature overnight.
  • the recrystallized THCA amine salt was then separated from the liquid phase by vacuum filtration, washed with 250mL cold heptane, filtered, and dried under vacuum to thereby yield 44.45 grams of a highly purified THCA-DMEA amine salt.
  • a sample of the 1x recrystallized salt was solubilized in methanol, diluted 250X and analyzed by HPLC (FIG. 7).
  • the solid primary THCA-DMEA amine salt was separated from the liquid phase by vacuum filtration, then re-slurried with 400 mL of cold heptane, vacuum filtered, and dried under vacuum to thereby yield 47.20 grams of a crystalline THCA-DMEA salt.
  • a sample of the crude THCA-DMEA salt was solubilized in Methanol, diluted 250X and analyzed by HPLC (FIG. 9).
  • the primary crystalline THCA-DMEA salt was then recrystallized by dissolving 36.20 g of THCA-DMEA salt in 3:1 volume/mass ratio ethyl acetate (109 mL) under refluxing conditions and then cooled slowly to 37 °C at which temperature the rate of precipitation of solid THCA amine salt increased rapidly after which, a 1 :1 volume ratio of warm heptane (109 mL, 30 °C) was added to the recrystallization mixture. The mixture was further cooled to 4 °C and then held at this temperature overnight.
  • the recrystallized THCA amine salt was then separated from the liquid phase by vacuum filtration, twice reslurried with 250 mL of cold heptane, filtered and dried under vacuum to thereby yield 33.193 grams of a highly purified THCA-DMEA amine salt.
  • a sample of the 1x recrystallized salt was dissolved in methanol, diluted 125X and analyzed by HPLC (FIG. 11 ).
  • a C. sativa biomass feedstock was processed with butane to produce a crude C. sativa extract in butane solvent.
  • the crude C. sativa extract was desolventized to thereby produce a dried C. sativa crude extract in a resin form.
  • the two washed and dried crude THCA amine salts were combined (2.993g) and re-slurried in 30mL cold heptane.
  • the solid THCA amine salts were then separated from the liquid phase by vacuum filtration and dried.
  • the washed and dried crude THCA amine salts were recrystallized by dissolution in in 8.6 mL ethyl acetate (3:1 vol/wt ratio) by heating the mixture to about 60 °C.
  • the reaction mixture was then cooled slowly to about 30 °C at which temperature the rate of precipitation of solid THCA amine salt increased rapidly and a 1 :1 volume ratio of warm heptane (8.6mL, 28°C) was added.
  • the mixture was further cooled to about 4 °C and held at this temperature for about 10 h.
  • the recrystallized THCA-DMEA amine salt was then separated from the liquid phase by vacuum filtration, washed with 30 mL cold heptane, filtered, and dried under vacuum to thereby yield 2.312 grams of a highly purified THCA-DMEA amine salt.
  • a sample of the 1x recrystallized salt was solubilized in methanol, diluted 25X and analyzed by HPLC (FIG. 15).
  • a standardized solution containing 102.78 mg/mL THCA in 1 -butanol (FIG. 17) was prepared by dissolving 1 .7958 grams of a reconstituted THCA solid in a 4.45:1 volume/mass ratio of 1 -butanol (8mL).
  • the reconstituted THCA solid was produced by (i) precipitating a THCA- DMEA crude salt from a standardized heptane extract solution, (ii) solubilizing the THCA-DMEA crude salt in dichloromethane (DCM), (iii) washing the solution of THCA-DMEA in DCM with 5% HCI to solubilize the amine into the aqueous layer, (iv) separating the organic layer from the aqueous layer in a separatory funnel, and (v) removing the DCM solvent from the organic layer by rotary evaporation to yield a crude THCA solid.
  • DCM dichloromethane
  • the two THCA-amine salts were each dissolved in HPLC-grade methanol and analyzed by HPLC.
  • the HPLC analysis of the THCA-DMEA amine salt is shown in FIG. 18.
  • the HPLC analysis of the THCA-1-piperidineethanol amine salt is shown in FIG. 19.
  • This example pertains to extraction of hemp biomass with 95% ethanol, then desolventizing the ethanolic crude complex extract to produce a concentrated crude complex extract, then resolubilizing the concentrated crude complex extract in heptane solvent from which a crude CBDA amine salt is precipitated.
  • the resolubilized crude hemp extract was further diluted with 3.3 L of heptane to produce a normalized crude hemp extract containing 110 mg/mL of CBDA.
  • the normalized crude extract was then spiked with 12.28% v/v denatured ethanol to thereby generate 20.96 L of a spiked normalized crude extract containing 90.24mg/mL of CBDA. 20.96L of the spiked normalized hemp extract in heptane spiked with
  • the washed and dried crude CBDA-TEA amine salt was recrystallized by dissolving 2854.38 grams of the CBDA-TEA salt in 10:1 volume/mass ratio of ethyl acetate (28.54L) spiked with 1.5% v/v heptane (428mL) under refluxing conditions in a 38L stainless jacketed reactor.
  • the reaction was heated with stirring from an overhead mixer until the temperature reached 76.1 °C.
  • the reaction mixture was then cooled slowly to 4 °C and held at this temperature overnight with continual stirring.
  • the recrystallized CBDA amine salt was then separated from the liquid phase by vacuum filtration, re-slurried in cold heptane, pressured filtered to separate the washed recrystallized CBDA amine salt from the liquid phase, and placed in a vacuum drying oven at 20 °C overnight, yielding 2,420 grams of a 1X recrystallized CBDA-TEA amine salt.
  • a sample of the recrystallized CBDA-TEA salt was solubilized in methanol, diluted 200X and analyzed by HPLC (FIG. 24).
  • a 38-L stainless jacketed reactor with distillation apparatus was equilibrated to 23 °C and placed under vacuum.
  • a 10:1 volume/mass ratio of a 2.5% Na2COs solution (12.2 L) was transferred into the reactor using vacuum.
  • the 2.5% Na2COs solution was degassed for 10 min while stirring with an overhead mixer.
  • the reactor was equilibrated to ambient pressure by the injection of N2 gas and the 2.5% Na2COs solution was sparged with a continuous flow of N2 gas for 10 min.
  • 1 ,210.9 g of a 1X recrystallized CBDA-TEA amine salt was transferred into the 38L stainless jacketed reactor while maintaining a slight positive pressure in the reactor by the sparging N2 gas flow.
  • the recrystallized CBDA-TEA amine salt was then mixed with the sparged 2.5% Na2COs solution and the reactor was placed under vacuum.
  • the reactor was equilibrated to ambient pressure by N2 gas injection and heated to 99.5 °C over a period of 8 h with periodic injections of N2 gas to aid the distillation of TEA released from the CBDA-TEA salt as the reaction proceeded.
  • the reaction mixture was then cooled to 71.3 °C and 4.8 L of heptane was transferred into the reactor using vacuum and mixed thoroughly for 30 min. The mixing was stopped and the biphasic solution was allowed to separate.
  • the upper organic layer was drained from the reactor and washed with 20 L of 5% HCI in a 30L glass carboy.
  • the organic layer containing decarboxylated CBD was separated from the aqueous layer containing the triethylamine-hydrochloride using a 20-L separatory funnel and collected in a 30-L glass vessel. The heptane liquid phase was then removed by distillation to yield 821.3 grams of a highly pure decarboxylated CBD oil. A sample of the decarboxylated CBD oil was solubilized in methanol, diluted 400X and analyzed by HPLC (Fig. 25).
  • This example pertains to extraction of hemp biomass with 95% ethanol, then desolventizing the ethanolic crude complex extract to produce a concentrated crude complex extract, then resolubilizing the concentrated crude complex extract in 1 -butanol solvent from which crude CBDA amine salts may be precipitated.
  • Dried hemp biomass was extracted with a 95% ethanol solvent at about 4 °C.
  • the extracted hemp biomass was separated from the ethanolic crude complex extract solution.
  • the ethanolic crude complex extract solution was desolventized to produce a crude complex extract oil.
  • 38.6 grams of the hemp crude complex extract oil was placed in an evaporator flask and any residual ethanol solvent was removed by distillation, resulting in 26.8 grams of a viscous crude resin.
  • a standardized extract solution containing 118.0 mg/mL of CBDA was prepared by dissolving the viscous crude resin in a 3:1 mass/volume ratio of 1 -butanol (80 mL) (FIG. 27).
  • FIGs. 27 to 30 for this Example show a very small peak at a retention time of 9 min that is identified as “THC-A”.
  • FIG. 31 shows an HPLC scan of an unfiltered HPLC-grade methanol control blank.
  • FIG. 32 shows an HPLC scan of a filtered HPLC-grade methanol control blank wherein an anomaly associated with the filter is confirmed but mis-identified as “THC-A” by the HPLC.
  • This example pertains to extraction of hemp biomass with heptane, then desolventizing the crude complex extract to produce a concentrated crude complex extract, then resolubilizing the concentrated crude complex extract in heptane solvent from which a crude CBDA amine salt was precipitated with triethylamine, washed, and recrystallized into a purified CBDA-triethylamine salt.
  • the purified CBDA-triethylamine salt was dissolved in dichloromethane (DCM) and washed with 5% HCI to precipitate CBDA, after which the DCM was remove by distillation to thereby yield a highly purified crystalline CBDA.
  • the CBDA was resolubilized in 1-butanol solvent after which nine selected amines were assessed for their ability to precipitate CBDA-amine salts from the CBDA-1- butanol solution.
  • a dried hemp biomass was extracted with heptane solvent.
  • the extracted hemp biomass was separated from the crude complex extract heptane solution.
  • the crude complex extract solution was desolventized to produce a crude complex resin extract.
  • the resin was resolubilized in heptane solvent and standardized after which, triethylamine (TEA) was added into the crude complex tor precipitate a crude CBDA-TEA amine salt.
  • TAA triethylamine
  • the crude CBDA-TEA amine salt was recovered, washed, then dried.
  • the washed CBDA-TEA amine salt was resolubilized in ethyl acetate, then recrystallized by addition of heptane as the antisolvent.
  • the purified recrystallized CBDA-TEA amine salt was recovered by vacuum filtration and then dried under vacuum.
  • the purified recrystallized CBDA-TEA amine salt was dissolved in dichloromethane (DCM) and then washed with a 5% HCI solution to solubilize the amine into an aqueous layer while the CBDA remained in the organic layer. After separation of the aqueous later, the DCM solvent was removed from the organic layer by distillation to thereby yield a solid crystalline CBDA product.
  • DCM dichloromethane
  • CBDA-TEA salt (FIG. 34), CBDA-HB salt (FIG. 35), CBDA-TBA salt (FIG .36), CBDA-DBN salt (FIG. 37), CBDA-DABCO (FIG. 38), and CBDA-1-piperidineethanol salt (FIG. 39) was dissolved in HPLC-grade methanol and analyzed by HPLC.
  • FIGs. 33 to 39 for Example 9 show a very small peak at a retention time of about 9 min that is identified as “THC-A. This peak is not due to the presence of THC-A but instead, is an unknown anomaly associated with the filters in the syringes used to prepare the samples for HPLC analyses (FIGs. 31 , 32).
  • This example pertains to extraction of hemp biomass with 95% ethanol, then desolventizing the ethanolic crude complex extract to produce a concentrated crude complex extract, then resolubilizing the concentrated crude complex extract in ethyl acetate solvent from which crude CBDA amine salts may be precipitated.
  • a standardized extract solution containing 94.694mg/mL of CBDA was prepared by dissolving the viscous crude resin in a 4:1 mass/volume ratio of ethyl acetate (169 mL) (FIG. 40).
  • TAA Triethylamine
  • DMEA dimethylaminoethanol
  • Hunig's base N,N- diisopropylethylamine
  • the solid amine-salts were separated from the liquid phase by vacuum filtration, washed with 10mL cold heptane and dried under vacuum.
  • a sample of each of the CBDA-TEA salt (FIG. 41 ), CBDA-DMEA salt (FIG. 42), and CBDA- Hunig’s base amine salt (FIG. 43) was dissolved in HPLC-grade methanol and analyzed by HPLC.
  • FIGs. 40 to 43 for Example 10 show a very small peak at a retention time of about 9 min that is identified as “THC-A. This peak is not due to the presence of THC-A but instead, is an unknown anomaly associated with the filters in the syringes used to prepare the samples for HPLC analyses (FIGs. 31 , 32).
  • EXAMPLE 11 EXAMPLE 11 :
  • This example pertains to extraction of hemp biomass with denatured ethanol, then desolventizing the ethanolic crude complex extract to produce a concentrated crude complex extract, then resolubilizing the concentrated crude complex extract in denatured ethanol solvent from which crude CBDA amine salts may be precipitated.
  • Dried hemp biomass was extracted with denatured ethanol (84.15% vol/vol ethanol, 15% v/v methanol, 0.85% v/v ethyl acetate) at -20 °C to produce an ethanolic crude complex extract.
  • the ethanolic crude complex extract. was desolventized by distillation to produce 23.4 g of a viscous concentrated crude complex resin.
  • the crude complex resin was resolubilized in 93.6 mL of denatured ethanol (4:1 vol/wt ratio) to produce a standardized standard complex containing 67.58 mg/mL CBDA (FIG. 44).
  • FIGs. 44 to 46 for Example 11 show a very small peak at a retention time of about 9 min that is identified as “THC-A. This peak is not due to the presence of THC-A but instead, is an unknown anomaly associated with the filters in the syringes used to prepare the samples for HPLC analyses (FIGs. 31 , 32).

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  • Crystallography & Structural Chemistry (AREA)
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Abstract

L'invention concerne des procédés de précipitation de sels de CBDA-amine et de sels de THCA-amine à partir d'extraits complexes bruts concentrés. Les extraits complexes bruts peuvent être récupérés à partir de biomasse de cannabis ou de systèmes de fermentation dans lesquels des micro-organismes ont été cultivés pour produire des cannabinoïdes cibles sélectionnés, à l'aide d'un solvant d'extraction sélectionné. Les extraits complexes bruts concentrés sont produits par déshydratation et désolvantation. Les extraits complexes bruts concentrés sont resolubilisés dans un solvant choisi parmi le méthanol, l'éthanol, l'éthanol dénaturé, le propanol, l'isopropanol, le butanol, le dichlorométhane, l'acétate d'éthyle et les alcanes en C5-C7. Certaines amines sont ajoutées aux extraits complexes bruts concentrés resolubilisés pour précipiter sélectivement des sels de CBDA-amine ou des sels de THCA-amine à partir de ceux-ci. Le sel de CBDA-amine précipité ou un sel de THCA-amine peut être purifié par recristallisation. L'amine peut être séparée du sel de CBDA-amine ou d'un sel de THCA-amine pour ainsi produire un CBDA ou THCA purifié. Le sel de CBDA-amine ou un sel de THCA-amine peut être décarboxylé pour produire un produit de CBD ou de THC.
PCT/CA2021/051745 2020-12-04 2021-12-06 Procédés de commutation de solvant pour la précipitation, le traitement et la purification de cannabinoïdes sélectionnés à partir de mélanges complexes concentrés Ceased WO2022115973A1 (fr)

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Cited By (1)

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US12054453B2 (en) 2019-05-17 2024-08-06 Mile High Labs, Inc. Systems and methods for refining cannabidiol

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CA2866787A1 (fr) * 2011-09-29 2013-04-04 Thc Pharm Gmbh The Health Concept Acides carboxyliques de cannabinoide, sels d'acides carboxyliques de cannabinoide, et fabrication et utilisation desdits acides et sels d'acides carboxyliques de cannabinoide
WO2019071213A1 (fr) * 2017-10-05 2019-04-11 Receptor Life Sciences, Inc. Formulations de cannabinoïdes synthétiques et à base de plante, à effet rapide et à action prolongée
WO2020016875A1 (fr) * 2018-07-19 2020-01-23 Al&Am Pharmachem Ltd. Procédé de purification d'acides cannabinoïdes à partir d'un extrait de matériel végétal
CA3111964A1 (fr) * 2019-06-12 2020-12-17 Nectar Health Sciences Inc. Procedes d'extraction, de traitement et de purification de famille selectionnee de composes cibles issus du cannabis

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CA2866787A1 (fr) * 2011-09-29 2013-04-04 Thc Pharm Gmbh The Health Concept Acides carboxyliques de cannabinoide, sels d'acides carboxyliques de cannabinoide, et fabrication et utilisation desdits acides et sels d'acides carboxyliques de cannabinoide
WO2019071213A1 (fr) * 2017-10-05 2019-04-11 Receptor Life Sciences, Inc. Formulations de cannabinoïdes synthétiques et à base de plante, à effet rapide et à action prolongée
WO2020016875A1 (fr) * 2018-07-19 2020-01-23 Al&Am Pharmachem Ltd. Procédé de purification d'acides cannabinoïdes à partir d'un extrait de matériel végétal
CA3111964A1 (fr) * 2019-06-12 2020-12-17 Nectar Health Sciences Inc. Procedes d'extraction, de traitement et de purification de famille selectionnee de composes cibles issus du cannabis
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US12054453B2 (en) 2019-05-17 2024-08-06 Mile High Labs, Inc. Systems and methods for refining cannabidiol

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