WO2021161147A1

WO2021161147A1 - A method of extraction for plants belonging to the cannabaceae family

Info

Publication number: WO2021161147A1
Application number: PCT/IB2021/050999
Authority: WO
Inventors: Kell Thomas; Michael LUMBY SØRENSEN; Patrick BAHRT GREGERSEN; Frans William LANGKILDE
Original assignee: Cibid A/S
Priority date: 2020-02-10
Filing date: 2021-02-08
Publication date: 2021-08-19

Abstract

Methods for the treatment of plants of the Cannabaceae family are disclosed by which the plants are treated with a combination of reduced pressure and sound waves. The isolated components resulting from the treatment may be used to formulate medicaments and other compositions, which are valuable tools and a major step forward in the management, the treatment or the alleviation of a number of diseases, conditions and disorders.

Description

A METHOD OF EXTRACTION FOR PLANTS BELONGING TO THE CANNABACEAE FAMILY

Field of the invention

5 The present invention generally relates to methods of treating plants belonging to the Cannabaceae family so as to isolate components of the Cannabaceae plant family. In particular, the components may be used for medical and/or nutritional purposes.

Ί0 Background of the invention

The Cannabaceae plant family is a small family of flowering plants. The family includes about 170 species grouped in about 11 genera, including Cannabis (hemp, marijuana), Humulus (hops) and Celtis (hackberries). The plants have many constituents which possess various properties.

15

Cannabinoids, terpenes (terpenoids) and other components of plants isolated from the Cannabaceae plant family are becoming more important as they have been shown to possess potent properties in the treatment or alleviation of various diseases and conditions.

20

The plants of the Cannabaceae family contain many compounds, reportedly up to approximately 400 different compounds according to some sources. Among them, approximately 85 known terpenoid-derived molecules are found only in cannabis plant variants and denoted cannabinoids. Although having similar 25 structure, they possess very different properties. Many of the cannabinoids are found in both acid and neutral forms and may have distinct medical properties.

Furthermore, approximately 200 terpene compounds and derivatives have been extracted from Cannabaceae plants. Terpenes are also found in other 30 plants, and as terpenes are generally volatile compounds, they are often associated with the characteristic smell and taste of the plant in question such as cineol (tea tree), citronellol (rose), limonene (citrus), and green tea (phytol) to mention a few. The components of the Cannabaceae plants act on certain receptors. Cannabinoid receptors are located throughout the body and form part of the endocannabinoid system. The endocannabinoid system is involved in a variety of physiological processes including appetite, pain-sensation, mood and memory. Cannabinoid receptors are a class of cell membrane receptors forming part of the G protein-coupled receptor superfamily. Cannabinoid receptors are activated by three major groups of ligands, namely endocannabinoids produced by the mammalian body, plant cannabinoids produced by the cannabis plant, and synthetic cannabinoids, respectively. The endocannabinoids and plant cannabinoids are lipophilic and, thus, fat-soluble compounds.

The cannabinoid receptors are mainly expressed in the brain (in particular the central nervous system (CNS)), but are also expressed in the lungs, the liver and the kidneys. The currently most recognised cannabinoid receptors are CBi and CB2, but other non-CBi and non-CB2 receptors expressed in endothelial cells and the CNS also exist.

Cannabinoids as such are a class of diverse chemical compounds acting on cannabinoid receptors by influencing and altering the neurotransmitter release.

The oils from the cannabis plant may comprise up to and possibly exceeding 400 components according to some sources, several of which components exhibit biological activity.

Thus, the potential of cannabinoids and terpenes is obvious.

In particular, terpenes also play a role in a number of nutritional products. Furthermore, terpenes possess various pharmacological properties, thereby contributing to the pharmacological effects attributed to other compounds present in the Cannabaceae plant. The observed effects may very well be due to intrinsic terpene properties, or the effects may be a result of a synergistic interaction between cannabinoids and terpenes. Some cannabinoids and terpenes may further produce reactants between a cannabinoid and a terpene. The use of extracted cannabinoids from plant material is controversial due to the fact that some of the components of the cannabis plant are classified as dangerous narcotics. This applies in particular to THC (delta9-tetrahydro- cannabinol) and derivatives thereof, whereas the other main constituent, CBD (cannabidiol) and derivatives thereof, are linked to other biological activity. Thus, in many countries, cannabinoids are prohibited as medicaments, unless the content of the narcotic components is below a certain value.

However, some of the THC derivatives cannot be excluded as potentially valuable for medicinal uses.

Therefore, it is considered important to obtain the components that do not possess the narcotic properties as well as those components showing the strongest beneficial potential. This has been tried in several ways, including specific extraction and isolation of various components, selective breeding of highly potent plants with a certain dominant profile, and synthesis of artificial cannabinoids.

E.g. from WO 2015 065544 A1 and WO 2014 145490 Al, plant varieties are known producing high contents of THC or THC derivatives, e.g. tetra- hydrocannabivarin (THCV), or CBD and related compounds. Higher content of the interesting cannabinoids eases extraction and isolation since the starting material has higher level of the desired components.

Conventionally used extraction methods include grinding plant material (such as leaves, stems, flowers, or seeds), drying the material, mixing the dried material with a solvent, heating and extracting oils. From e.g. WO 2003 064847 Al, a method for extracting cannabis plants is known. The method includes mixing finely ground cannabis material with methanol and chloroform as extraction solvent, applying ultrasound for 15 minutes, decanting the solution and subjecting the solution to chromatography. The obtained resulting herbal drug extract is then mixed with methanol and subjected to chromatography for further purification. Other conventional extraction methods are described by e.g. Journal of pharmaceutical and Biomedical Analysis 143, 228-236 (2017), and US 2012 0059062. The conventional methods include maceration, percolation and extraction with solvents like alcohol and liquid carbon dioxide of herbal material. The so obtained extract may be further purified by heating or chilling. US 2013 0079531 describes methods for extraction of active ingredients from plant material using a cold solvent and very short mixing/contact period. Freshly harvested or dried plant material is used in the extraction, and stems, stalks, seeds as other material apart from leaves and flowering buds are removed. During this short period of time, the herbal material may be shaken or treated with low-intensity vibrations, after which the solvent is separated quickly through pressing, centrifugal screening, or expulsion. The total process takes 30 seconds.

Several other extraction methods are known.

E.g. CN 106 831 353 describes a method, whereby the Cannabis material is crushed and dried at 60-200°C, followed by extracting the dried material with ethanol, and subsequently drying the material to obtain a dried powder. The extraction step may be performed using reflux extraction, ultrasonic extraction and/or soak extraction on the dried powder. Extraction on the dried powder may be performed at 60 Hz, or 70 Hz in 70% ethanol.

US 2013/0251824 describes methods for the extraction of cannabinoids, by which method a hopper is filled with plant material, a solvent for extraction is transferred from a solvent tank, and the solvent containing the extract is recovered. The method may be supplemented with soaking, washing or rinsing as this facilitates extraction of cannabinoids into the solvent. The washing procedure may be performed under stirring such as mechanical stirring or ultrasonic stirring. Furthermore, the residual plant material may be dried using a vacuum pump or preferably a solvent pump to evaporate remaining solvent.

From US 2019/0241536, methods or the extraction and concentration of cannabinoids are known. Shredded freshly harvested plant material is mixed with solvent to form a slurry and is subjected to ultrasound treatment to release intracellular contents of the plant material into the solvent. Subsequent filtering treatment is applied to remove residual plant material, waxes and chlorophyll, whereafter the solvent is subjected to vacuum distillation to evaporate the solvent from the cannabinoids.

US 2019/0134531 describes extraction of cannabis using ultralow temperature extraction. The ultralow temperature is reached using liquid nitrogen under pressure and controlled by means of heat exchangers.

From US 9,718,065, a process for trichome separation is known. The separation process comprises agitating, chopping or grinding (e.g. physical agitation) trichome-bearing plant material, separating predetermined size fractions of trichomes and plant material, and removing the floating fraction of trichomes followed by rinsing the trichome fraction. The plant material may be agitated in an ice bath in a wide mesh sieve bag (preferably 220 micron) to release the trichomes into the ice and water slurry optionally containing e.g. ice, ceramic beads, or chemical compounds. The plant material contains also cell walls and nuclei and needs further processing.

In LJS 2005/0165088, cannabis extracts rich in CBD, CBDA, CBDV and CBDVA substances, preferably for the treatment of nausea, vomiting, emesis, motion sickness and the like are described. The extracts are obtained from plant material such as bark, wood, leaves, stems, roots, flowers, fruits, seeds, or berries. The extracts are prepared by maceration, percolation, solvent extraction or treatment with carbon dioxide of the plant material. Further purification may be performed by supercritical or subcritical extraction, vaporisation and chromatography. The resultant extract contains non-specific lipid-soluble material which can be removed by chilling to -20°C followed by filtration.

US 8,445,034 discloses systems and methods for fabricating a medicine from cannabis plant material. The methods allow selective utilisation of various cannabinoid molecules from whole cannabis sativa. The whole cannabis sativa plant may, after drying or heating, be treated with home-based extractors (such as reflux or ultrasonic) to produce extracts that are unique combinations of medicinal phytocannabinoids enabling patients to titrate oral cannabis tinctures. It is specified that extraction is not effective unless the plant material is dried and thereafter ground. To preserve the acid form of the desired cannabinoids, ultrasonic or reflux extraction is performed below 200°F (approximately 94°C), preferably between 140°F and 175°F (approximately 60°C to 80°C). The frequency of the home-based ultrasonic apparatus is not defined.

US 2011 0256245 discloses methods for obtaining a cannabis plant extract suitable for medical uses. The method comprises obtaining cannabis flower trimmings with trichome material, covering the material with cold water, agitating the mixture, soaking the cannabis flower trimmings in cold water, removing the cannabis trimmings and removing the trichome material from the water, followed by drying of the trichome material. The dried material may be formulated into a paste to an ointment or may be used to create a paste/cream.

US 6,243,896 relates to in situ manufacturing of plant extracts from living plants. The extract is manufactured from blooms, buds, leaves, stalks, seed, berries, and aerial roots. The plant is not uprooted and remains rooted during the extraction process, i.e. the plant is maintained in its natural surroundings. In certain embodiments, a portion of the flower or other component of the plant is brought into contact with an extraction solution for a period of time. The extraction is performed using a vessel, a cup or a bowl (preferably made from beeswax) holding the extraction solution and placed around the e.g. plant flower. The extraction period is between three hours and several weeks and takes place at ambient temperature. Examples of suitable plants are mentioned, however, no trichome-bearing plants are mentioned specifically.

WO 2000 07437 relates to methods of identifying and recovering products exuded/secreted of plants. In particular, biologically active compounds are exuded/secreted from the cuticle of a plant (the leaf surface). By the method, the surface is brought into contact with a solvent dissolving the cuticular material, the solvent containing the cuticular material is assayed, and the cuticular material is analysed for agents with biological activity. The method may be performed on living plants. The method of collecting the cuticular material is performed at ambient temperature.

One major disadvantage by the conventionally used methods is that it is difficult to obtain the various components of the cannabis plants in an easy and cost-effective manner. Accordingly, there is a need for improved extraction methods.

Summary of the invention

By the present invention, improved methods of extracting components from plants belonging to the Cannabaceae plant family are provided. The components may be used to formulate e.g. medicaments, nutritional products and other compositions, which are valuable tools and a major step forward in the management, the treatment or the alleviation of a number of diseases, conditions and disorders. Thereby, the real potential of cannabinoids, including terpenes, can be exploited.

In a first aspect, the present invention provides a simple and easy to perform way to extract components such as oils from plants belonging to the Cannabaceae family. By said method, the Cannabaceae plants are placed in a vessel, and a solvent is supplied to the vessel, after which a combination of reduced pressure and sound waves is applied to the mixture of solvent and Cannabaceae plants.

Accordingly, the present invention relates to a method for the treatment of plants belonging to the Cannabaceae family comprising the steps of:

- placing the plants of the Cannabaceae family in a vessel,

- supplying a solvent to the vessel,

- applying a reduced pressure below atmospheric pressure, and

- applying sound waves, while maintaining the reduced pressure.

The reduced pressure facilitates removal of air bubbles from the plants and plant parts so as to ensure better contact between plants/plant parts and solvent. Thereby, a more effective extraction is obtained. Furthermore, it is advantageous to use a reduced pressure, since the reduced oxygen may result in less oxidation of cannabinoids and terpenes present. Some solvents tend to thicken at lower temperatures, and, therefore, the reduced pressure improves the flowability of the solvent.

The plants applied in the method of the present invention may be freshly harvested or dried, ground, chopped or whole plants or plant parts. The plant parts may comprise stems, stalks, seeds as other material in addition to leaves and flowering buds.

By applying a reduced pressure and sound waves, while maintaining the reduced pressure, components of the Cannabaceae plant material are effectively released into the solvent. Subsequently, the solvent with the valuable components can be withdrawn from the vessel and subjected to further purification (e.g. filtering, crystallisation, winterization and/or HPLC) to isolate the desired components.

The isolated components may be used for preparing medical compositions for the treatment, management or alleviation of a number of diseases, disorders or conditions. Such diseases, conditions and disorders include, but are not limited to, cancerous diseases, chemotherapy-induced nausea and vomiting, auto-immune diseases such as psoriasis, colitis ulcerosa and Crohn's Disease, and neurological disorders such as epilepsy, spasticity associated with Multiple Sclerosis or spinal cord injury, sleep disorders, depression, and pain relief e.g. associated with chronic pain as well as other diseases.

The isolated components may further be used for preparing nutritional compositions or formulations, or dietary products for alleviating certain conditions or to be taken as supplement to a healthy diet.

Medical and nutritional compositions may further comprise other substances such as, but not limited to, carriers, adjuvants, and excipients such as e.g. solvents or solubilising agents, diluents, buffers, suspending agents, wetting agents, pH-adjusting agents, fillers, dispersing agents, preserving agents, flavouring agents and colorants. The medical and nutritional compositions may take several forms including, but not limited to, pills, tablets, chewing gums, liquids, solids, powders, creams, gels, and suppositories. In particular in the case of nutritional compositions, nutritional formulations and dietary products, such may suitably also be formulated as bars, chewables or drinks for oral ingestion.

The amount of the isolated components and the chemical nature of the isolated components depend on the specific application and administration routine. Methods for preparing medical compositions and nutritional compositions/ - dietary products involve procedures generally known in the art for formulating such.

Brief description of the drawings The invention is further illustrated by the accompanying drawings. The drawings are for illustrative purposes only and are non-limiting on the scope of the invention. In the drawings,

Fig. 1 shows extraction by the method of the invention (Example 1),

Fig. 2 shows extraction by a conventional extraction method (Example 3),

Fig. 3 shows a comparison between plant material extracted according to the invention and according to the conventional methods (Examples 1 and 3),

Fig. 4 shows the analysis of the extract obtained in Example 1 following evaporation of the solvent, and

Fig. 5 shows the analysis of the further purified extract prepared according to Example 4.

Detailed description of the invention The various aspects of the present invention are described in more detail in the following.

Several plant species have developed trichomes which have a variety of functions depending on the plant species. Trichomes on plants are epidermal outgrowths of various kinds. A common type of trichomes is hairy outgrowths and may be unicellular or multicellular, branched or unbranched. In particular, multicellular hairs may have one or several layers of cells. Branched trichomes (or hairs) may be dendritic, i.e. tree-like, or stellate, i.e. star-shaped. Another common type of trichomes is peltate hairs having a plate or shield-shaped cluster of cells attached directly to the surface or borne on a stalk. Any of the types of trichomes may be glandular, meaning that they produce a secrete such as oils. As mentioned, the trichomes are very variable in their presence among different plant species and may be located on different plant organs and vary in size, density and functionality. The functionality i.a. interferes with feeding of herbivores or protects the plant surface cells from frost, windy conditions, or even sunlight. In some cases, the trichomes are a defence system for the plant, i.e. like the stinging hairs of nettles.

Cannabis plants may be either female or male, or in some, although rare, cases both. Various differences exist among the female and male plants. Most notable are the large resin-secreting flowers produced by the female plants as well as the calyxes containing high concentrations of trichomes that secrete cannabinoids including CBD, THC, and terpenes. Furthermore, glands located on the buds secrete i.a. terpenes. Thus, extraction of these oils or resins is essential in order to obtain components having therapeutic properties.

The present invention relates to a method for the treatment of plants of the Cannabacaeae plant family using a combination of reduced pressure and sound waves.

Thus, the present invention relates to a method for the treatment of plants of the Cannabaceae plant family, comprising the steps of

- placing the plants of the Cannabaceae family in a vessel,

- supplying a solvent to the vessel,

- establishing a reduced pressure in the vessel,

- applying sound waves, and

- applying the sound waves, while maintaining the reduced pressure. The present method provides a method of extracting components from plants of the Cannabacaeae plant family, by which method the extract comprises less undesired components such as waxes, fats and the like.

The plants of the Cannabaceae family are placed in a vessel having one or more valves, the vessel and the valves being sealable and connectable to, e.g., a pump, such as a vacuum pump, and a sound wave generator. The vessel should further be such which can withstand the reduced pressure.

Following the combined treatment with reduced pressure and sound waves, the solvent is recovered from the vessel. The solvent may suitably be recovered from the vessel by means of a pump, e.g. a solvent pump or a vacuum pump. The solvent comprises the components from the Cannabacaeae plants released during the reduced pressure/sound wave treatment. The solvent may be poured from, sucked from e.g. using a pump, or in other ways separated from the residual plant material. The solvent may be collected in another vessel, a jar, a container and the like. The solvent may subsequently be subjected to further treatment to isolate the extracted components (e.g., HPLC, solvent evaporation, washing, and/or winterization).

Within the present context, the term "plants" is intended to include freshly harvested or dried, ground, chopped or whole plants or plant parts. The plant parts may comprise stems, stalks, seeds as other material in addition to leaves, flowers and flowering buds. In some embodiments, the plants specifically include trichomes.

Within the present context, the term "reduced pressure" is intended to mean a pressure below atmospheric pressure and may be expressed either as a percentage of the atmospheric pressure or may be expressed in another conventionally used unit, such as e.g. "bar" or "Pascal". In general, atmospheric air pressure is given in millibars (mbar), where standard atmospheric pressure at sea level is defined as 1,013.25 mbar. Representative examples of reduced pressures to be used in the method of the invention include, but are not limited to, 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% and 100%, respectively, atmospheric pressure as well as any integer or non-integer values therebetween.

In one embodiment of the method according to the invention, the reduced pressure is between 0 and 1000 mbar. Other examples of a suitable reduced pressure are 10 mbar, 20 mbar, 30 mbar, 40 mbar, 50 mbar, 100 mbar, 200 mbar, 300 mbar, 400 mbar, 500 mbar, 600 mbar, 700 mbar, 800 mbar and 900 mbar as well as any integer and non-integer value therebetween. In particular, the reduced pressure may be between 10 mbar and 750 mbar. In some embodiments, the reduced pressure may be between 10 mbar and 500 mbar. Specifically, the reduced pressure may be between 10 mbar and 250 mbar.

Thereby, air bubbles are effectively removed from the plants/plant parts, and oxidation of the cannabinoids and terpenes are further minimised due to the reduced oxygen content. The reduced pressure also facilitates better distribution of the solvent around the plant material. This is in particular an advantage if a thick viscosity solvent is used, such as e.g. ethanol which tends to thicken at lower temperatures.

The method of the present invention further comprises applying sound waves, while maintaining the reduced pressure. Thereby, material and components of the plant material are released into the solvent.

In general, sound is defined as a vibration that propagates in a transmission medium such as a liquid, a gas or a solid. In the art, sound waves are differentiated by their frequencies, and, thus, acoustic sound waves are such with frequencies from 20 Hz to 18 kHz, and ultrasound waves are such with frequencies from 18 kHz to 18 MHz.

For the purpose of the present invention, sound waves having a frequency of from 18 kHz to 1 MHz are applied.

Thereby, the sound waves further facilitate the removal of air bubbles from the plants/plant parts, whereby better contact between the solvent and the plants/plant parts is obtained. Furthermore, the sound waves may further facilitate extraction of various oils and resins, in particular from the trichomes as the sound waves are capable of "knocking off" the trichomes.

Within the context of the invention, a suitable solvent is such, which can receive the components of the Cannabaceae plants or plant material. The components need not be dissolved in the solvent as long as the solvent is able to function as a carrier or emulsifier for droplets of the components. Suitable solvents include, but are not limited to, water, organic solvents, and mixtures of water and organic solvents. Organic solvents include, but are not limited to, alcohols, alkanes and ethers as well as mixtures thereof. Specific examples of organic solvents include alcohols such as methanol, ethanol, and propanol, alkanes such as n-hexane, pentane and cyclohexane, and ethers such as petroleum ether, di-isopropyl ether, and dioxane. Other suitable organic solvents include, but are not limited to, toluene, formic acid, chloroform, acetaldehyde, acetone and acetonitrile. Suitable mixtures of water and organic solvent may be 80% w/w (weight percent) water to 20% w/w organic solvent (indicated as 80:20). Other suitable mixtures include water to organic solvent in the ratios 99: 1, 95:5, 90: 10, 85: 15, 80:20, 75:25, 70:30, 65:35, 60:40, 50:50, 40:60, 30:70, 20:80, 10:90, or 1:99 (all % w/w). In particular, mixtures of water and alcohol such as ethanol may be 99: 1, 95:5, 90:10, 85: 15, 80:20, 75:25, 70:30, 65:35, 60:40, 50:50, 40:60, 30:70, 20:80, 10:90, or 1:99 (% w/w). It is to be understood that the organic solvent may be a mixture of different organic solvents in the range of 1-99 % w/w of one organic solvent to another organic solvent. By way of example, a mixture of ethanol to methanol may be 10:90, 20:80, or 30:70 (all % w/w). In particular, the solvent may be a mixture of water and ethanol, or water in combination with ethanol and methanol in the beforementioned ratios. Water may be any type of water such as tap water, however, it may be preferred that the water is purified or ion exchanged water, such as e.g. water sold under the tradename "Ultra Clean Water". The solvent may further comprise other components, e.g. to adjust the ionic strength, to obtain a buffer effect or capacity, to adjust the pH, or to form salts. Such other components include, but are not limited to, salts e.g. sodium chloride, sodium bicarbonate and ammonium formate. In one embodiment of the method according to the invention, the solvent is a 20:80 % w/w mixture of water and ethanol. The water may preferably be Ultra Clean Water. Usually, the ethanol used is commercially available 96% ethanol.

Within the context of this invention, the suitable temperature of the solvent may be in the range of from -80°C to 25°C, or from -20°C to 5°C, such as from -5°C to 2°C, or 1°C. It is to be understood that the temperature may in particular depend on the period of time of applying reduced pressure and sound waves. The temperature should preferably be such at which the solvent does not boil at the selected reduced pressure. The solvent may suitably be precooled to the desired temperature in a vessel equipped with cooling means and transferred to the vessel containing the plants of the Cannabaceae family by means of a pump, e.g. a solvent pump or vacuum pump.

Thus, in one embodiment of the method according to the invention, the temperature of the solvent is -80°C, -2°C, 0°C, 1°C or 25°C.

Within the context of this invention, the suitable period applying reduced pressure and sound waves is in the range of from 1 second to 60 minutes. Specific examples include, but are not limited to, 1, 10, 20, and 30 seconds, 1 minute, 2 minutes, 5 minutes, 10 minutes, 15 minutes, 20 minutes, and 30 minutes. Additional mechanical stirring may suitably be applied.

As mentioned above, sound waves within the present context are defined as having frequencies from 18 kHz to 1 MHz. In general, within the purpose of this invention, the sound wave frequency will be such that does not damage the plant cells, e.g. by permeabilising or sonicating the plant cells. In particular, the frequency may be from 18 kHz to 120 kHz. It is to be understood that the frequency may in particular be 18 kHz, 20 kHz, 30 kHz, 40 kHz, 50 kHz, 60 kHz, 70 kHz, 80 kHz, 90 kHz, 100 kHz, 110 kHz, or 120 kHz as well as any integer or non-integer value therebetween. In particular, a frequency of 20, 30 or 40 kHz may be used. Furthermore, different frequencies may be used during the application of the sound waves. The frequency will depend on the type of plant and its content of oils (oil composition or components), the temperature of the solvent, the nature of the solvent as well as the period of time of applying sound waves.

The application of sound waves with lower frequencies (below or at maximum 1 MHz) is advantageous, since it does not permeabilise or sonicate the plant or plant material due to lower operating frequencies as does conventionally used ultrasound having frequencies above 1 MHz. Thus, the plant or plant material is preserved much more efficiently, and it is easier to separate plant material from the solvent after treatment with reduced pressure and sound waves.

By applying sound waves, the components are extracted from the plants without destroying the plant cells unnecessarily. Thereby, it is easier to separate the solvent from the plant material for further purification or separation of desired components.

Also, the procedure will be influenced by the time of application of reduced pressure and sound waves, the reduced pressure, the frequency of the sound waves, the solvent, and the period of time.

For illustration purposes, the method of the invention may suitably be carried out the following way:

- placing the plants of the Cannabaceae family in a vessel having one or more valves, at least one of the valves being connected to a vacuum pump,

- supplying a solvent, the solvent optionally being supplied to the vessel from another vessel equipped with cooling means to precool the solvent to a desired temperature,

- establishing a reduced pressure below the atmospheric pressure in the vessel with the plants of the Cannabaceae family using the vacuum pump,

- closing the one or more valves of the vessel with the plants of the Cannabaceae family to maintain the reduced pressure,

- applying sound waves to the vessel with the plants of the Cannabaceae family by means of a sound wave generator. The vessel with the plants of the Cannabaceae family may be equipped with one or more valves, one of the valves being connected to a vacuum pump, and another valve for safety purposes. The (extraction) vessel with the plants of the Cannabaceae family may further be equipped with cooling means or may be isolated to maintain a desired temperature during extraction The solvent may suitably be supplied to the vessel with the plants of the Cannabaceae family from another vessel equipped with cooling means for precooling the solvent to a desired temperature and a valve for connection to a pump. The solvent may suitably be supplied using a pump connected both to the vessel with the plants and vessel with the solvent, and the solvent is transferred from the solvent vessel to the plant vessel. The pump used for transferring the solvent may be the same as the pump for establishing the reduced pressure (a vacuum pump) or another pump (e.g., a solvent pump). After transfer of the solvent to the vessel with the plants of the Cannabaceae family, the valve to the solvent vessel is closed. The vessel with the plants of the Cannabaceae family is sealed, and the vacuum pump is used to establish the desired reduced pressure. When the desired reduced pressure is reached, the valve to the vacuum pump is closed and the vacuum pump is switched off. The reduced pressure will be maintained effectively. Thereafter, sound waves are applied for a suitable period of time as specified above. The temperature of the solvent may be as specified above. It is desired that the temperature of the solvent is such that the solvent does not boil at the selected reduced pressure. After a certain period with reduced pressure sound wave treatment, the sound wave generator is switched off. The solvent now comprising the extracted components of the Cannabacae plants is recovered from the vessel, e.g., by starting the vacuum pump (or a solvent pump) and opening the valve connecting the vacuum pump to the vessel, or by a hose connected to the valve since the reduced pressure will result in the flow of the solvent when the valve is opened. If cold solvent is used, a vacuum pujjp is used to transfer the solvent with the extracted components. A standard solvent pump is generally not preferred as cold solvent will damage a standard solvent pump.

In one embodiment of the method according to the invention, the solvent comprising the extracted components of the Cannabaceae plants is further subjected to separation by filtering. The filtering may be accomplished using one or more filters having a well-defined size (mesh). The filtering may be performed in order to separate impurities, e.g., plant residues, to remove components, e.g. proteins. The filtering may be performed once in one step or through a series of one or more filters. The filtering may also be performed by re-circulating the mixture of the oil-containing fraction and the solvent repeatedly a number of times through one or more filters. The temperature of the solvent during such further filtering treatment is as defined above.

In some embodiments of the method of the invention, the solvent comprising the extracted components may be subjected to a centrifugation step prior to the filtering step. In other embodiments of the invention, a centrifugation step may be performed after the extraction, but before subsequent steps.

Within the context of this invention, the plants belonging to Cannabaceae family may be selected from cannabis species. Suitable strains and varieties include, but are not limited to, Cannabis sativa, Cannabis indica and Cannabis ruderalis, and hybrids thereof as well as genetically modified cannabis plants. In particular, the strains and varieties may be used in combination to benefit from the intrinsic composition of components produced by the individual species. The method of the invention is also very suited in connection with industrial cannabis plants (certain strains of Cannabis sativa and Cannabis ruderalis).

In certain embodiments of the invention, it may be advantageous to use cannabis plants of a specific phenotype. Cannabis plants usually have a content of THC of around 12-25%, while the CBD level is below 4%. Cannabis strains producing CBD of more than 4% are considered high CBD strains. Cannabis strains producing CBD in a content of 20% have emerged in the past few years. However, it is understood that no cannabinoids are produced during the vegetative cycle. Cannabinoids are produced during the flowering period. Thus, in one embodiment of the invention, the cannabis species are selected so as to be such high CBD strains (high-CBD cannabis plants). In another embodiment of the invention, the cannabis species are selected so as to be such high THC strains (high-THC cannabis plants). In other embodiments, the cannabis species are selected so as to be high-terpenes strains.

Cannabis plants are in general known to produce cannabinoids such as CBGA, which transform to CBG, CBDA, THCA and all other minor cannabinoids, followed by further transformation, into CBD and THC to mention a few. The synthesis of the components during the plants' lifecycle/season is well- described in the art. Depending on which components are desired, the harvesting and separation methods may be chosen having regard to the particular cannabis plants and the cannabis plant profile.

The method of the invention may further comprise a filtering step. Thereby, unwanted plant material may be removed from the obtained extract. Thereafter, the extract can be subjected to further processing. The filtering step may be performed using one or more filters. During the filtering process, a series of filters may suitably be selected having mesh sizes of from 250 pm to 25 pm, such as e.g. 250 pm, 175 pm, 150 pm, 125 pm, 115 pm, 100 pm, 75 pm, 50 pm, 45 pm, and 25 pm. In one embodiment, four filters are used having the mesh sizes 250 pm, 175 pm, 50 pm, and 25 pm. Filters are generally commercially available, and a broad selection of filtering sizes and varieties exists.

The solvent with the extracted components may be subjected to further purification, e.g., winterization, crystallisation or preparative HPLC or may be used directly for other purposes, e.g., for the preparation of a medicament, a supplement or dietary product. Winterization is a well-known procedure and may be used to remove fats and lipids from the solvent, yielding a residue with a higher purity with regard to cannabinoids. Subsequently, the solvent may be subjected to evaporation to separate solvent and desired components (crude oil with the cannabinoids). However, in particular at lower solvent temperatures, winterization may not be needed since no waxes or fats are contained in the solvent.

As mentioned, the solvent with the extracted components of the Cannabaceae plants may be subjected to further separation or purification by high- performance liquid chromatography (HPLC), preferably preparative HPLC, or crystallisation in suitable solvents such as pentane. HPLC and preparative HPLC procedures as well as crystallisation procedures are generally well- described in the art. However, the conditions when used in connection with the present invention have to be adapted in view of the composition of the desired components.

Cannabis plants contain many compounds, reportedly up to approximately 400 different compounds according to some sources, among them approximately 85 known terpenoid-derived molecules which are found only in cannabis plants and denoted cannabinoids. Although having similar structure, they possess very different properties. The cannabinoids are found predominantly in the trichomes of the female cannabis plants. Many of the cannabinoids are found in both acid and neutral forms and may have distinct medical properties.

Some of the most notable cannabinoids are:

CBG-A Cannabigerolic acid

CBG Cannabigerol

THCA Tetrahydrocannabinolic acid

THC-C4 Tetrahydrocannabinol-C4

THCVA Tetrahydrocannabivarinic acid

A9-THC Delta-9-tetrahydrocannabinol

A8-THC Delta-8-tetrahydrocannabinol

THCV Tetrahydrocannabivarin

CBNA Cannabinolic acid

CBN Cannabinol (primary product of THC degradation)

CBDA Cannabidiolic acid

CBDVA Cannabidivaric acid

CBD Cannabidiol

CBDV Cannabidivarin

CBCA Cannabichromic acid

CBC Cannabichromene

CBLA Cannabicyclol acid Apart from those cannabinoids found intrinsically in the cannabis plants, ageing or curing (exposure to e.g. heat, light and air) may be used to change the profile of the constituents of the plant flowers. Thereby, other canna binoids may be obtained, many of which have medical activity. E.g. CBDA and THCA may be modified by decarboxylation. Some with possible medical activity are listed below.

As evident from the above table, the cannabinoids found in the cannabis plant potentially possess many important properties. However, it is also evident that, for medicinal applications, it is important to obtain only those cannabinoids that do not show the narcotic side effects, or alternatively at least reduce the contents of cannabinoids with narcotic side effects. For dietary or food products, the content of certain cannabinoids as well as other components, for this application, may be different from those of medical applications.

A number of important terpenes with proposed pharmacological properties exist. These include the following:

Many of the other cannabinoids can probably be expected to be of medicinal/pharmacological importance, and in particular some of the minor components may support or enhance the effect of others. In particular, the terpenes interact with the cannabinoids to create additional or enhanced effects (the so-called "entourage effect").

Typically desired components of the Cannabaceae plants obtained according to the methods described herein comprise either

1) THC and derivatives thereof, e.g. THCA, as well as minor components, 2) CBD and related compounds, e.g. CBDA and CBG, as well as minor components,

3) minor cannabinoids, i.e. those other than THC, THCA, CBN, CBD, CBDA, and CBG, or

4) a number of terpenoids, e.g. those mentioned above in the table.

Different components may suitably be mixed to obtain a suitable overall composition of the components, i.e. for application as medicament, supplement or dietary product.

The cannabis plant contains many other components apart from the cannabinoids, namely terpenoids (many known), hydrocarbons (50 known), nitrogen-containing compounds (more than 70 known), carbohydrates (13 monosacharides, 2 disacharides, 5 polysacharides), flavonoids (23 known), fatty acids (33 known, mainly unsaturated), non-cannabinoid phenols (34 known) as well as a number of simple alcohols, aldehydes, ketones, esters and lactones. All of these may have some functions together with the cannabinoids.

In another embodiment, it is desirable that components obtained by the present method are composed of less than 1-10% THC, THCA and other THC derivatives. In some embodiments, it is desirable that the components obtained by the present method are composed of less than 2%, 3%, 4%, 5%, 6%, 7%, 8% or 9% (as well as any non-integer therebetween) THC, THCA and/or other THC derivatives.

In one embodiment, it is desirable that the components obtained by the present method are composed of more than 50% cannabidiol (CBD), preferably 60-70% CBD. In some embodiments, the obtained components are composed of up to 60%, 61%, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69% or 70% (as well as any non-integer therebetween). In some embodiments, it is desirable the extracted components are composed of 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% (as well as any non-integer therebetween) CBD. In one embodiment, it is desirable that the components obtained by the present method are composed of more than 90% THC. In some embodiments, the obtained components are composed of up to 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% (as well as any non-integer therebetween) THC.

As mentioned above, it may be an advantage to mix different components so as to obtain a composition with a specific content of cannabinoids.

Analytic procedures and methods for determining the presence of various constituents/components are e.g. described in Monograph Cannabis Flos Version 7.1 (November 28, 2014).

The components obtained by the method as defined herein are useful for preparing a medical composition for the treatment or alleviation of a number of diseases, disorders or conditions. Such include, but are not limited to, cancerous diseases, chemotherapy-induced nausea and vomiting, auto immune diseases such as psoriasis, colitis ulcerosa and Crohn's Disease, and neurological disorders such as epilepsy, spasticity associated with Multiple Sclerosis or spinal cord injury, sleep disorders, depression, and pain relief e.g. associated with chronic pain.

The medical composition may be administered in any suitable way. Such include administration by smoking, vaporisation, oral ingestion (e.g. pills, powders, tablets, liquids, gels), transdermal application (e.g. creams, oils, gels, ointments), intravenous injection, intraarticular injection, subcutaneous injection, intramuscular injection, sublingual absorption (e.g. pills, tablets, oils), infusion, or rectal suppository.

The medical composition of the invention may further include other substances such as, but not limited to, carriers, adjuvants, and excipients such as e.g. solvents or solubilising agents, diluents, buffers, suspending agents, wetting agents, pH-adjusting agents, fillers, dispersing agents, preserving agents and colorants. The medical composition of the invention may take several forms including, but not limited to, pills, tablets, liquids, solids, powders, creams, gels, ointments, suppositories and formulations for smoking/vaporising. Specific suitable carriers, adjuvants, and excipients are generally well-known in the art.

The medical composition may suitably be administered once or several times per day, week or month, depending on the disease, disorder or condition to be treated or alleviated, and the dose contained in the medicament. In some cases, the medical composition may be administered for prophylactic purposes.

Nutritional preparations may take several forms including, but not limited to, drinks, chewables, bars, pills, tablets, solids, and powders. Nutritional preparations may further include e.g. vitamins, minerals, fluids, colorants, flavouring agents, fillers, and proteins, to mention a few. The nutritional preparations may suitably be administered once or several times per day, week or month, depending on the disease, disorder or condition to be treated or alleviated, and the dose contained in the nutritional preparation. In some cases, the nutritional preparation may be administered for prophylactic purposes.

The invention is further illustrated by the non-limiting examples.

Examples Example 1

Treatment of plants of the Cannabaceae plant family using the method of the invention.

In this example, a total of 3 stainless steel vessels was used. The dimensions of the vessels were 50x30x15cm, and each contained 22 I.

Equipment and plants were kept at a temperature of 5°C in order to keep the process stable.

Vessel 1. Pre-Cool vessel (placed in freezer at -80°C)

Vessel 2. Extraction vessel. Vessel 3. Product vessel.

All vessels were supplied with a depressurizing safety valve, opening when the vacuum drops below 50 mbar. The safety valves were adjustable and adjusted to the selected vacuum for the process.

The Pre-Cool vessel was filled with solvent (20 I, 96% ethanol, freezing point approximately -117°C) and sealed off. Subsequently, the vessel was cooled to -80°C.

2000 g of plant material, in this example industrial hemp tops (Cannabis sativa var. Finola) was used. The plant material was ground, using a shredder/grinder. Ground plant material was transferred to an extraction bag.

The extraction bag was made to fit the extraction vessel, generally in 90-115 pm polyester/nylon. In this example a 115 pm bag was used, due to a coarser grind. The extraction bag was placed in the extraction vessel, then the vessel was sealed off.

The extraction vessel was depressurised to 50 mbar using a vacuum pump. By slowly opening a valve the -80°C ethanol from Pre-Cool vessel was supplied to the extraction vessel by a pipe connected to the extraction vessel, until the Pre-Cool vessel was empty, and the extraction vessel was full (i.e. the solvent was transferred). After supplying all ethanol solvent from the Pre-Cool vessel to the extraction vessel, the Pre-Cool vessel was closed off using the valve on the pipe connecting the Pre-Cool vessel and extraction vessel. The process took less than 1 minute, but obviously depends on the amount of solvent transferred.

The vacuum was kept at 50 mbar during the whole process, to remove air bubbles in plant material. After the vacuum of 50 mbar was reached, sound waves having a frequency of 30 kHz was applied for 5 minutes, while maintaining the vacuum at 50 mbar. The vacuum pump was kept switched on in case the vacuum was not at 50 mbar at all times. Thereby, cannabinoids were extracted from the plant material. Thereafter, the sound wave treatment was stopped, and subsequently the vacuum was discontinued. Now, the "Product vessel" was depressurised, and when vacuum was at 100 mbar, the valve, from the Extraction vessel to the Product vessel was slowly opened. Thereby, the extracted solution was drawn into the Product vessel. Then, the solution in the Product vessel was hand filtered at 25 pm. Alternatively, a filter could be inserted between the Extraction vessel and the Product vessel to ease the filtering process.

The following was observed: The method of the invention performed excellent in terms of a better and faster extraction of the plants. No chlorophylls or unwanted fat or lipids got extracted (visual inspection). In comparison with conventional ethanol extractions methods, the method of the invention extracted the plants faster. Furthermore, it is believed that the low temperature during the extraction contributes to the pure extract observed. Also, the transfer from the Extraction vessel to the Product vessel at low temperature is believed to influence the purity of the extract.

The solvent with the extracted components is shown in Fig. 1 and Fig. 3 (right). As clearly seen in Fig. 1 and Fig. 3 (right), the method of the invention yielded a nice and clear solution, indicating that no (or only minimal) impurities are present in the solvent. It was concluded that the combined use of vacuum (reduced pressure) and sound wave treatment resulted in an extract with an excellent content of Cannabis components (having regard to the components produced by the Cannabis plants which of course is dependent on the Cannabis strain used). The use of sound waves knocks off the oils located on the trichomes without damaging the plant material, and, thus, the extract does not contain significant amounts of, e.g., chlorophyll and other unwanted substances which need to be removed subsequently. Accordingly, the method of the invention makes possible large-scale production of pure products since the sound waves can easily penetrate large amount of plant material and facilitate extraction of cannabinoids.

Analysis of the extracted components is shown in Fig. 4. The analysis was performed on the oil obtained after evaporation (standard procedure) of the ethanol solvent. The analysis verified that the method of the invention captures the wanted components quickly and efficient.

Example 2

We changed the vacuum for the extraction, from 50 mbar to 500 mbar.

The vacuum to draw over the ethanol to the product vessel, was kept the same, at 100 mbar.

Also, the temperature of the ethanol was still kept at -80°C.

Vessel 1. Pre-Cool vessel (placed in freezer at -80°C)

Vessel 2. Extraction vessel.

Vessel 3. Product vessel.

All vessels were supplied with a depressurizing safety valve, opening when the vacuum drops below 500 mbar. The safety valves were adjustable and adjusted to the selected vacuum for the process.

The Pre-Cool vessel was filled with solvent (20 I, 96% ethanol) and sealed off. Subsequently, the vessel was cooled to -80°C.

2000 g of plant material, in this example industrial hemp tops (Cannabis sativa var. Finola) was used. The plant material was ground, using a shredder/grinder. Ground plant material was transferred to an extraction bag. The extraction bag was made to fit the extraction vessel, generally in 90-115 mih polyester/nylon. In this example a 115 pm bag was used, due to a coarser grind. The extraction bag was placed in the extraction vessel, then the vessel was sealed off.

The extraction vessel was depressurised to 500 mbar, and the vacuum was kept, while transferring the solvent from the Pre-Cool vessel. By slowly opening a valve the -80°C ethanol from Pre-Cool vessel was supplied to the extraction vessel by a pipe connected to the extraction vessel.

After supplying all ethanol solvent from the Pre-Cool vessel to the extraction vessel, the Pre-Cool vessel was closed off using the valve on the pipe connecting the Pre-Cool vessel and extraction vessel. The process took about 5 minutes, but obviously depends on the amount of solvent transferred.

It was observed that due to the vacuum (500 mbar) being closer to atmospheric pressure, it took almost 5 times longer to transfer the solvent from the Pre-Cool vessel.

The vacuum was kept at 500 mbar during the whole process, to remove air bubbles in plant material. After the vacuum of 500 mbar was reached, sound waves having a frequency of 30 kHz was applied for 5 minutes, while maintaining the vacuum at 500 mbar.

Thereby, cannabinoids were extracted from the plant material. Thereafter, the sound wave treatment was stopped, and subsequently the vacuum was discontinued. Now, the "product vessel" was depressurised, and when vacuum was at 100 mbar, the valve, from the Extraction vessel to the Product vessel was slowly opened. Thereby, the extracted solution was drawn into the Product vessel. Then, the solution in the Product vessel was hand filtered at 25 pm. Alternatively, a filter could be inserted between the Extraction vessel and the Product vessel to ease the filtering process.

The following was observed: The method of the invention performed excellent in terms of a better and faster extraction of the plants. No chlorophylls or unwanted fat or lipids got extracted (visual inspection). Using a reduced pressure of 500 mbar during the extraction, all the desired cannabinoids got into the solution.

In comparison with conventional ethanol extractions methods, the method of the invention extracted the plants faster. Furthermore, it is believed that the low temperature during the extraction contributes favourably to the pure extract observed. Also, the transfer from the Extraction vessel to the Product vessel at the used low temperature is believed to influence the purity of the resulting extract.

Example 3

Prior art extraction for comparison

For comparison with the method of the invention, a conventional extraction ("bucket tech" extraction) was performed:

2000 g industrial hemp tops (Cannabis var. Finola) was used. The plant material was ground, using a shredder/grinder. Ground plant material was transferred to a 22 I vessel and filled with 20 I 96% ethanol, cooled to 5°C. The vessel was sealed off and stirred. The vessel was left for approximately 15 minutes. Subsequently, the solvent was hand filtered using a 115 pm filter to separate plant material from the extract. The resulting solvent with the extracted components is shown in Fig. 2 and Fig. 3 (left). As it can clearly be seen, the conventional extraction method yields a very dark (almost black) extract, indicating the heavy presence of impurities, making further purification absolutely necessary. In comparison, the extract obtained by the present method (cf. Fig. 3 (right)) is clear and without discolouration.

Example 4

Isolation of CBD from the extract prepared according to Example 1

The extract of Example 1 was further purified by evaporation (standard procedure). The various cannabinoids ware separated using the inherent difference between evaporation temperature of different cannabinoids. In this Example, CBD from the crude oil (the oil from Example 1 after evaporation of ethanol solvent) was separated and collected.

Analysis of the purified product is shown in Fig. 5. As can be seen, the product consists of 99% CBD. Thus, the oil can be considered ultrapure. THC as well as THCA are not detected in the purified oil, thus, making the oil very useful for applications where THC and THCA are unwanted components.

It was concluded that the method of the invention makes possible easy further processing of extracts resulting in pure cannabis products having a desired content of specific cannabinoids. No harsh chemicals or conditions were needed for purifying the starting material (the crude oil of Example 1). This is important if the cannabis extract is to be used in medicinal or nutritional products as any residual solvents or components are not allowed due to human health and/or environmental issues.

Claims

1. A method for the treatment of plants of the Cannabaceae plant family, comprising the steps of

- placing the plants of the Cannabaceae family in a vessel,

- supplying a solvent to the vessel,

- establishing a reduced pressure in the vessel,

- applying sound waves, and characterised in that the method comprises

- maintaining the reduced pressure, while applying sound waves.

2. A method according to claim 1, characterised in that the reduced pressure is between 0 and 1000 mbar.

3. A method according to claim 1 or 2, characterised in that the reduced pressure is between 10 mbar, 20 mbar, 30 mbar, 40 mbar, 50 mbar, 100 mbar, 200 mbar, 300 mbar, 400 mbar, 500 mbar, 600 mbar, 700 mbar, 800 mbar and 900 mbar as well as any integer and non-integer value therebetween.

4. A method according to any one of the preceding claims, characterised in that the sound waves have a frequency of from 18 kHz to 1 MHz.

5. A method according to any one of the preceding claims, characterised in that the solvent is a mixture of water and an organic solvent.

6. A method according to any one of the preceding claims, characterised in that the Cannabaceae plant family are selected from Cannabis sativa, Cannabis indica and Cannabis ruderalis, hybrids or genetically modified variants thereof, or a combination thereof.

7. A method according to any one of the preceding claims, characterised in that the plants are industrial Cannabis plants such as Cannabis var. Finola.

8. A method according to any one of the preceding claims, characterised in that the plants are Cannabis plants producing more than 4% CBD, or Cannabis plants producing 12-26% THC.

9. A method according to any one of the preceding claims, characterised in that the temperature during applying reduced pressure and sound waves is from -20°C to 5°C, or -80°C to 25°C.

10. A method according to any one of the preceding claims, characterised in that the method further comprises the steps of: centrifugation to separate solvent and plants of the Cannabaceae plant family and, filtering.