Abstract
A method for the extraction and isolation of the terpene and isoprenoid compounds from plant material, followed by a centrifugal force induced selective crystallization of isoprenoids resulting in a separation of terpene and isoprenoid fractions. This this method is suitable for the extraction of cannabinoids from Cannabis and the enrichment tetrahydrocannabinolic acid and reduction of tetrahydrocannabinol in an extract. The purity of tetrahydrocannabinolic acid resulting from centrifugal crystallization is such that dissolution and selective recrystallization of tetrahydrocannabinolic acid is possible resulting in >99.9% pure tetrahydrocannabinolic acid, w/w.
Claims
1. A method of extracting terpene and isoprenoid compounds from plant material and for selectively enriching the concentration of isoprenoid compounds in a fraction of this extract comprising: a. Cannabis plant material into an extraction chamber; b. degassing the extraction chamber; c. introducing chilled solvent into the extraction chamber; d. filtration of the extract-containing solvent; e. removal of the extract-containing solvent from the extraction chamber; f. placing the extract-containing solvent into a centrifuge vessel; g. centrifuging the centrifuge vessel to promote crystallization and to separate the crystals from the supernatant solvent; and h. collecting the THCA containing crystals.
2. The method of claim 1 wherein the solvent is subcritical n-propane, butane, hexane, pentane, ether, ethyl acetate, heptane, toluene, naphtha, methanol, ethanol, isopropanol, butanol, or combinations thereof.
3. The method of claim 1 wherein the solvent is supercritical carbon dioxide.
4. The method of claim 1 wherein various extracts of Cannabis are substituted for the plant material.
5. The method of claim 1 further comprising addition of sugar, salt, or other precipitants to the extract containing solvent prior to centrifugation.
6. The method of claim 1 further comprising the collection of the terpene rich fraction.
7. A method of increasing the purity of THCA in partially purified THCA containing material comprising: a. dissolving THCA containing material into a solvent; b. transferring a portion of THCA containing solvent into a container; c. initiating THCA crystallization; d. adding of additional THCA containing solvent as THCA crystals form; e. separating THCA crystals from residual solvent; and f. collecting the THCA crystals.
8. The method of claim 7 wherein the solvent is subcritical n-propane, butane, hexane, pentane, ether, ethyl acetate, heptane, toluene, naphtha, methanol, ethanol, isopropanol, butanol, or combinations thereof.
9. The method of claim 7 wherein the solvent is supercritical carbon dioxide.
10. The method of claim 7 further comprising addition of acid, base, or buffering compounds to the solvent or solvent containing extract to regulate the pH of the system and to promote differential crystallization.
11. The method of claim 7 wherein crystallization is initiated by evaporating off some solvent.
12. The method of claim 7 wherein crystallization is initiated by reducing the temperature of the system.
13. The method of claim 7 wherein crystallization is initiated by seeding with sugar or cationic salts, cationic liquids, or THCA crystals.
14. The method of claim 7 wherein THCA crystals are washed with additional solvent prior to collection to remove impurities left in the residual solvent used in initial dissolution of the partially purified THCA containing material.
15. The method of claim 1 further comprising inert gas sparged into the extraction chamber, collection vessel, or centrifuge vessel prior to solvent extraction of the plant material to minimize oxidation of THCA.
16. The method of claim 7 further comprising dissolution of partially purified THCA material into solvent taking place under inert gas and with inert gas sparged solvent and the recrystallization of THCA taking place under inert gas.
17. The method of claim 1 further comprising addition of hydrophilic reducing agent to the plant material prior to extraction to minimize oxidation of THCA.
18. The method of claim 1 further comprising addition of hydrophobic reducing agent to the solvent prior to plant material extraction to minimize oxidation of THCA.
19. The method of claim 7 further comprising addition of hydrophobic reducing agent to the dissolution and recrystallization solvent to minimize oxidation of THCA.
20. The method of claim 1 further comprising addition of pH buffering agent to the plant material prior to extraction to minimize oxidation of THCA.
21. The method of claim 7 further comprising saponification of recrystallized THCA.
22. The method of claim 21 further comprising crystallization of the THCA salt.
23. The method of claim 7 further comprising application of a coating to the THCA crystals.
24. The method of claim 23 wherein this coating is oxygen impermeable.
Description
BRIEF DESCRIPTION OF THE FIGURES
[0023] FIG. 1a shows the chemical structure of the non-psychoactive compound tetrahydrocannabinolic acid (THCA).
[0024] FIG. 1b shows the chemical structure of tetrahydrocannabinol (THC).
[0025] FIG. 2 is a table showing academic studies that have shown medicinal effects of THCA, and the stability of THCA in vivo.
[0026] FIG. 3 shows the decarboxylation reaction by which TCHA is degraded into THC.
[0027] FIG. 4a is a box diagram showing the pre-fractionation extraction process by which terpene and isoprenoid compounds are extracted from Cannabis, similar to that shown in the primary embodiment of U.S. Provisional Application Ser. No. 62/146,198.
[0028] FIG. 4b is a box diagram showing the post-extraction fractionation process by which isoprenoid and terpene compounds are separated from Cannabis extracts, similar to that shown in the primary embodiment of U.S. Provisional Application Ser. No. 62/146,198.
[0029] FIG. 5a is a box diagram showing the primary embodiment of this invention.
[0030] FIG. 5b is a table showing one example of primary embodiment, with this table presenting data on the enrichment of THCA from plant biomass to enriched extract through use of the primary embodiment of this invention, with THCA values shown having been measured by mass spectrometry (MS) following separation by high pressure liquid chromatography.
[0031] FIG. 6 is a box diagram showing the second embodiment of this invention.
[0032] FIG. 7 is a box diagram showing the third embodiment of this invention.
[0033] FIG. 8 is a box diagram showing the fourth embodiment of this invention.
[0034] FIG. 9 is a box diagram showing the fifth embodiment of this invention.
[0035] FIG. 10a is a box diagram showing the sixth embodiment of this invention.
[0036] FIG. 10b shows the chemical structure of THCA salts formed in the sixth embodiment of this invention, with X.sup.+ preferably being from the group consisting of NH.sub.4.sup.+, mono-, di- or trivalent metal ions, and primary, secondary, tertiary or quaternary organic ammonium ions with up to 48 C atoms, which may bear still further functional groups.
[0037] FIG. 11 is a box diagram showing the seventh embodiment of this invention.
DETAILED DESCRIPTION OF THE INVENTION
[0038] With reference to FIG. 4a and FIG. 4b, the initial purification of isoprenoid compounds from Cannabis is conducted similarly to the process previously described in U.S. Provisional Application Ser. No. 62/146,198 entitled A Method for Extracting Cannabinoids and Terpenes through Centrifugation, filed Apr. 10, 2015. FIG. 4a shows a flow chart representing the initial extraction process by which THCA, as well as other cannabanoids, isoprenoids, and terpenes are extracted from Cannabis. In 301, the female flowers of Cannabis are harvested and destemmed. In 302 the destemmed flowers from 301 are shredded, macerated, ground, milled, or otherwise reduced in size. In 303, the macerated flower material from 302 is chilled. In 304, the chilled flower material from 303 is placed into a pre-chilled extraction chamber. In 305, the extraction chamber is sealed and degassed. In 306, supercritical (liquid) n-propane is introduced into the degassed extraction chamber from 305, with this n-propane dissolving and extracting chemical compounds from the flower material. In 307, the liquid n-propane containing dissolved compounds extracted from the flower material from 306 is separated from the flower material solids by filtration. In 308, the filtered liquid n-propane containing dissolved compounds extracted from the flower material from 307 is collected into a receiver vessel. In 309, the n-propane is removed from the extract contained in the receiver vessel in 308. In 310, the extract-containing receiver vessel, from which n-propane had been recovered in 309, is removed from the extraction chamber apparatus, and this raw extract is ready for further fractionation.
[0039] In some embodiments, the flowers are not dried. In one preferred embodiment, the flowers are reduced in size to 1-3 mm. In some embodiments, the flowers are not reduced in size. In one preferred embodiment, the flowers are chilled to between 0 C. and 100 C. prior to extraction. In one preferred embodiments the extraction chamber is pre-chilled to between 0 C. and 100 C. In one preferred embodiment, the n-propane is pre-chilled to between 0 C. and 100 C. before introduction into the extraction chamber. In some embodiments, there is a soak time upon introduction of the n-propane into the extraction chamber prior to separating the n-propane containing dissolved compounds from the plant material. In other embodiments there is no soak time, and separation of the n-propane containing dissolved compounds from the plant material is immediate. In some embodiments, the n-propane is recovered from the receiver vessel by use of differential pressure or temperature such that the n-propane becomes a gas and is readily removable, as is known in the art.
[0040] FIG. 4b shows a flow chart representing an initial THCA purification step following the extraction process from U.S. Provisional Application Ser. No. 62/146,198 entitled A Method for Extracting Cannabinoids and Terpenes through Centrifugation, filed Apr. 10, 2015.In 311, the raw extract removed from the receiver vessel 310 (from FIG. 4a) it taken for fractionation. In 401, the raw extract from 311 is placed on a surface. In 402, the raw extract on a surface from 401 is placed in a vacuum oven, with this oven then being closed and purged of air. The extract is incubated in the vacuum oven until polymorphic crystals begin to form, at which point it is removed, 403. In 404, the extract removed from the vacuum oven containing polymorphic crystals from 403 is placed into a temperature controlled centrifuge, with the centrifuge then being activated so as to separate the solid crystals from the liquid phase of the extract. In 405, the centrifugally pelleted crystal fraction and liquid supernatant fraction from 404 are separated. In 406, the crystals from 405, which contain isoprenoid compounds crystals, are collected. In 407, the liquid supernatant fraction from 405, which is rich in terpenes, is filtered to remove any residual crystals. In 408, the filtered terpene-rich extract from 407 is recovered.
[0041] In some embodiments, the process may further comprise steps of filtering the condensate during centrifugation to collect water insoluble material and extracting the insoluble material. In some embodiments precipitants are added to the extract, in order to affect the precipitation of impurities, which will then be subsequently filtered out. In some embodiments precipitants are added to the extract in order to affect the selective precipitation of target cannabinoid compounds, with subsequent centrifugation or filtration being used to harvest these target compounds. In one preferred embodiment, sucrose crystals (sugar) can be used as a selective precipitant for THCA relative to other cannabinoids found in a non-polar solvent extract of cannabinoidswith this selectivity owing to the hydrophilic nature of sucrose and the relatively greater hydrophilic nature of THCA to other cannabinoids (resulting from the carboxylic acid moiety on THCA).
[0042] The primary embodiment of this invention is shown in FIG. 5a. In 501, isoprenoid compound crystals from 406 are dissolved into hexane. In 502, a portion of the hexane solution of dissolved compounds is poured into a crystallization chamber. In 503, a portion of the hexane or other solvent is evaporated while maintaining low temperature, by methods including but not limited to pulling a vacuum or blowing dry nitrogen gas on the solution. In 504, additional hexane solution containing dissolved compounds is added to the crystallization chamber as THCA crystals form and/or as solvent is evaporated. In 505, THCA crystals are removed from the residual THCA-depleted solution. In 506, THCA crystals are briefly washed in a small volume of nonpolar solvent to remove residual THCA-depleted solution and any contamination compounds that may have adsorbed to the external surface of the THCA crystals. In 507, washed THCA crystals are separated from the wash solution. In 508, THCA crystals are collected and stored. In some embodiments, crystals are stored under inert gas and/or in UV resistant containers, such as packaging THCA crystals in argon filled amber glass ampules. In some embodiments, a nonpolar solvent other than hexane is used, such as pentane, heptane, butane, ether, or any of a host of other solvents known in the art, or mixtures of specific solvents. In some embodiments, the initial THCA containing material is extracted and somewhat purified by a method other than that described in FIG. 4a and FIG. 4b, though we note that unless the initial THCA purity level is rather high (>90% THCA), subsequent re-crystallization of THCA may be impossible or impractically slow.
[0043] In some embodiments, the organic solvents used for recrystallization may include but are not limited to butane, propane, ethyl acetate, heptanes, toluene, ethanol, methanol, isopropanol and combinations thereof. In certain advantageous embodiments the target compound(s)/solvent mixtures temperature is brought to between 10 Celsius and 70 Celsius and precipitates are filtered out. In certain advantageous embodiments the organic solvent comprises ethyl acetate. In these and other embodiments it may also be preferable to adjust pH of the extract/solvent mixture to enhance precipitation of at least one target compound(s).
[0044] As an example of the primary embodiment, consider the preparation of a highly pure THCA Cannabis extract meant for use in research into the potential treatment of a human disease state, such as prostate cancer or various inflammatory diseases. (THCA has been indicated as potentially useful in treating these disease states, as in known in the art, with some examples being shown in FIG. 3.) While medicinal benefits have been shown for THCA, the narcotic effects of delta-9 THC and sedative effects cannabidiol (CBD) are undesirable in medicinal applications. Through use of the method of the primary embodiment, the purity of THCA from a sample of Cannabis can be greatly increased, possibly to within specifications for pharmaceutical usage (>99.9% purity) with an effective elimination of delta-9 THC and CBD, with this material having no intoxicating effect on the patient being treated. An example of the enrichment of THCA through use of the method of the primary embodiment is shown as a table in FIG. 5b. Briefly, 100 g of dried, mature Cannabis flower, with an initial THCA content of 17.8% w/w, subject to extraction by the method of U.S. Provisional Application Ser. No. 62/146,198 using subcritical n-propane as a solvent, with the initial extract having 49.5% THCA w/w, and the centrifugally separated crystals being 94.5% THCA w/w. Through use of the primary embodiment of this invention the purity of THCA in the re-crystalized material was increased to 99.97% with only limited THCA yield loss relative to the centrifugally separated crystals (30.3% and 31.3% yield, respectively, as shown in FIG. 5b). THCA analysis was done using an AGILENT 1100 (Agilent, Santa Clara, Calif.) HPLC with G1946D mass spectrometry detector using methods known in the art of liquid chromatography coupled mass spectrometry.
[0045] The second embodiment of this invention is shown in FIG. 6, in which the extraction and crystallization process of material from Cannabis takes place in chambers and solvents that are sparged with inert gas in order to remove oxygen from the process and prevent the degradation of THCA to THC. Destemmed, macerated Cannabis is placed in the extraction chamber in 304. Inert gas is then applied to the extraction chamber in 600, and this inert gas is subsequently removed by vacuum degassing in 601. In 602, inert gas is bubbled through liquefied n-propane. In 603, the liquid n-propane is separated from the headspace gas (including both sparge gas and any gas driven from the liquid n-propane, including H.sub.2S, O.sub.2, or CO.sub.2). In 604, the sparged n-propane is stored under inert gas. In 605, the sparged n-propane from 604 is applied to the degassed extraction chamber and macerated Cannabis from 601. In 607, the Cannabis extract in liquid n-propane is filtered from the residual plant material under inert gas. In 608, the filtered extract is placed into an inert gas sparged collection vessel. In 609, the initial crystallization of isoprenoid compounds, including centrifugation as previously described in this application, takes place under inert gas. In 610, isoprenoid compound crystals are separated under inert gas. In 611, isoprenoid compound crystals are dissolved into inert gas sparged hexane. In 612, THCA crystals are formed under inert gas. In 613, THCA crystals are stored under inert gas. In some embodiments, the inert gas is argon, nitrogen, or other inert gas known in the art.
[0046] As an example of the second embodiment of this invention, consider the extraction of THCA from Cannabis as an initial step in pharmaceutical production. As THCA is degraded to THC by oxygen, the removal of oxygen from the system by inert gas sparging increases the final THCA yield. As THCA is degraded by acid, the removal of H.sub.2S and CO.sub.2, both of which result in acid formation with dissolution in water (the small amount of water in biomass absorbs these gases, particularly at low temperatures), results in improved THCA yield. In addition to improving yield, in some applications, even trace THC contamination is restrictive. Through use of the second embodiment of this invention, THCA yield is increased and THC contamination is avoided.
[0047] The third embodiment of this invention is shown in FIG. 7, in which reducing agents are added to various points in the extraction and crystallization process to prevent THCA oxidation to THC by removal of dissolved oxygen from solvents or systems. Hydrophobic reducing agents, 700, can be added to n-propane, 701, or hexane (or other nonpolar re-crystallization solvent), 702. Hydrophilic reducing agent, 703, can be added to harvested Cannabis flowers 704, macerated flowers, 705, or crystal wash solution, 706. Hydrophobic and hydrophilic reducing agents of various compositions are known in the art.
[0048] As an example of the third embodiment of this invention, consider the extraction of THCA from Cannabis as an initial step in pharmaceutical production. As THCA is degraded to THC by oxygen, the removal of dissolved oxygen from the system by use of reducing agents (that react with dissolved oxygen and oxygen species) increases the final THCA yield. Through use of the third embodiment of this invention, and introducing reducing agents into one or more steps in the process, THCA yield is increased and THC contamination is eliminated.
[0049] The fourth embodiment of this invention is shown in FIG. 8, in which buffering, basic, or pH regulating compounds are integrated into the process in order to minimize the degradation of THCA. In 800, pH regulating agents are introduced into the nonpolar elements of the system in order to remove acid or acid-forming gasses (i.e., CO.sub.2 or H.sub.2S), with these compounds from 800 being used for solvent treatment or gas scrubbing of n-propane, 803, n-propane extracts of Cannibis, 801, or re-dissolved THCA crystals in hexane, 802. In 804, pH buffering agents that are water soluble are added to harvested flowers, 805, macerated flowers, 806, or crystal wash solution, 807. In some embodiments, the nonpolar solvents or gasses are treated with bases or gas scrubbing agents known in the art. In some embodiments, the pH buffering agents include any of the organic or inorganic pH buffering agents known in the art.
[0050] As an example of the fourth embodiment of this invention, consider the extraction of THCA from Cannabis as an initial step in pharmaceutical production. As THCA is degraded to THC by acid, the removal of dissolved acidic compounds or acid-forming gasses from the system by use of base or pH buffering agents increases the final THCA yield. Through use of the fourth embodiment of this invention, and introducing pH regulating or acid removing compounds into one or more steps in the process, THCA yield is increased and THC contamination is eliminated.
[0051] The fifth embodiment of this invention is shown in FIG. 9, in which cationic compounds are added to initiate or accelerate the crystallization of THCA from nonpolar solvent. Cationic compounds, 908, can be introduced into the hexane solution of dissolved isoprenoid compounds, 501, or into the crystallization chamber, 502. In some embodiments, the cations are introduced as salts. In some embodiments, cations are introduced as ionic liquids. In some embodiments, the cation is monovalent. In some embodiments, the cationic compound is multivalent.
[0052] As an example of the fifth embodiment of this invention, consider the industrial purification of THCA from extract. By making use of the fifth embodiment of this invention, the rate of crystallization can be increased, reducing labor usage as well as allowing for less installed tank/infrastructure per unit product.
[0053] The sixth embodiment of this invention is shown in FIG. 10a and FIG. 10b, in which the recrystallized THCA is reacted with base to form a THCA salt, with this THCA salt being of improved stability relative to THCA. Regarding FIG. 10a, in 1000, THCA crystals from 508 are re-dissolved into a solvent, such as an intermediate chain alcohol solution (e.g., 95% isopropanol). In 1001, base is then added to this semipolar solvent solution under controlled conditions in order to saponify the dissolved THCA. In 1002, the THCA-salt is crystallized and separated from the solvent and base. In 1003, the THCA-salt crystals are washed to remove any residual solvent and base. In 1004, THCA-salt crystals are collected and store. Regarding FIG. 10b, the general structure for a THCA-salt is shown, with X being the cation conjugate ion from the base. Suitable bases for the formation of crystalline salts are primary, secondary and tertiary organic amines with up to 48 carbon atoms such as dicyclohexylamine, ammonia, alkoxides, hydroxides, carbonates, hydrogen carbonates, carboxylates and other basic salts of elements of the first, second and third main group and of tin, lead and bismuth, and the alkoxides, hydroxides, carbonates, hydrogen carbonates, carboxylates and other basic salts of transition elements such as silver (Ag+). Inorganic salts may be complexed (e.g. silver hydroxide as silver diammine complex) in order to increase the solubility. Further suitable organic bases are pharmaceutical active substances with at least one basic nitrogen atom in the molecule, such as morphine, hydromorphone (Palladon), buprenorphine, etc. Suitable solvents depend on the base used, including semipolar solvents such as 95% isopropanol (5% water) and nonpolar solvents such as hexane, or other solvents known in the art suitable for saponification, including alcohols, esters, ethers, ketones, hydrocarbons, halogenated hydrocarbons and nitriles with up to 20 carbon atoms.
[0054] As an example of the sixth embodiment of this invention, consider the production of THCA for pharmaceutical usage in which the stability of THCA (and lack of THC contamination) is vital. Formation of a salt is widely utilized in the pharmaceutical industry as a way to impart stability to a compound. Through use of the sixth embodiment of this invention, THCA is stabilized to a THCA-salt. Saponification of cannabinoids from Cannabis has been previously shown in US 20150038567, however, prior methods resulted in mixed cannabinoid preparations including other cannabinoids, such as cannabidiol (CBD), which have different pharmacological effects than THCA and are undesirable in certain applications. In this invention, THCA is first purified from other isoprenoid compounds, including cannabinoids compounds such as CBD, prior to saponification of THCA to a salt.
[0055] The seventh embodiment of this invention is shown in FIG. 11, in which THCA crystals, 508, are coated with a material that is impermeable to water and acts as a barrier to UV light, 1100, prior to the packaging of the THCA crystals for storage or subsequent pharmaceutical use, 1101. In some embodiments, this coating is sprayed or tumbled onto crystals or compressed masses of crystals as seen in the pharmaceutical industry. In some embodiments, this coating is any of a plurality of compositions known in the art.
[0056] As an example of the seventh embodiment of this invention, consider THCA crystals being prepared for storage transit from an extraction facility to a pharmaceutical production facility. Through use of the seventh embodiment, the THCA could be stabilized in such a form as to allow this transportation without degradation of the THCA by air or UV light during transit and handling.
[0057] In some embodiments, the various embodiments can be combined. For example, extraction and recrystallization could take place under inert gas, and the final THCA crystals produced under inert gas could be coated with an impermeable layer.
[0058] Although described with reference to preferred embodiments of the invention, it should be recognized that various changes and/or modifications of the invention can be made without departing from the spirit and scope of the invention. In any case, the invention is only intended to be limited by the scope of the following claims.
