Patents: Synthetic Cannabidiol
MICROORGANISMS AND METHODS FOR THE FERMENTATION OF
CANNABINOIDS
WO2019071000
Disclosed herein are microorganism and methods that can be
used for the synthesis of cannabigerolic acid (CBGA) and
cannabinoids. The methods disclosed can be used to produce CBGA,
?9-tetrahydrocannabinolic acid (THCA), cannabidiolic acid
(CBDA), cannabichromenic acid (CBCA), ?9 -tetrahydrocannabinol
(THC), cannabidiol (CBD), cannabichromene (CBC). Enzymes useful
for the synthesis of CBGA and cannabinoids, include but are not
limited to acyl activating enzyme (AAE1), polyketide synthase
(PKS), olivetolic acid cyclase (OAC), prenyltransferase (PT),
THCA synthase (THCAS), CBDA synthase (CBDAS), CBCA synthase
(CBCAS), HMG-Co reductase (HMG1), and/or farnesyl pyrophosphate
synthetase (ERG20). The microorganisms can also have one or more
genes disrupted, such as gene that that controls beta oxidation
of long chain fatty acids.
METHOD FOR
PURIFYING CANNABINOID COMPOUNDS
CA3023760
The present invention relates to methods for purifying one
or two cannabinoid compounds using simulated moving bed
chromatography, wherein the cannabinoid compound(s) is/are
obtained in the extract and/or the raffinate with the total
amount of isomeric impurities being below detection level. In
particular, the present invention relates to methods for the
purification of cannabidiol,
trans-(-)-delta-9-tetrahydrocannabinol, cannabidivarin,
trans-(-)-delta-9-tetrahydrocannabivarin and cannabigerol which
have been obtained by enantiopure synthesis.
[0002] The present invention relates to methods for purifying
one or two cannabinoid compounds using simulated moving bed
chromatography, wherein the canna- binoid compound(s) is/are
obtained in the extract and/or the raffinate with the total
amount of isomeric impurities being below detection level. In
particular, the present invention relates to methods for the
purification of cannabidiol, trans-(-)-
delta-9-tetrahydrocannabinol, cannabidivarin, trans-(-)-
delta-9- tetrahydrocannabivarin and cannabigerol which have been
obtained by enantiopure synthesis. Furthermore, the present
invention also relates to an extract and/or raffinate which
is/are obtained or obtainable by the method accord- ing to the
invention. Since the discovery of the endogenous cannabinoid
system with its functional significance in terms of the
regulation and modulation of the immune as well as the nervous
system, there is an ongoing need for natural and artificial
canna- binoids for their selective, pharmaceutical control. In
particular, because of their different medical functions, there
is a need for targeted, separate stimulation of ao the
cannabinoid receptors CB1, which are mainly found in neurons, in
highest density in basal ganglia, in the hippocampus and the
cerebellum, and of the cannabinoid receptors CB2, which are
mainly found on cells of the immune sys- tem and on cells that
are involved in bone formation and bone loss. CA 03023760
2018-11-09 WO 2017/194173 PCT/EP2016/060905 - 2 - The
cannabinoid receptors CB1 and CB2 are presumed to be the
accepted sites of action of molecules with a cannabinoid
structure. Even though further recep- tors are discussed as
potential CB3 receptors, it is assumed that the main effects are
mediated via CB1 and CB2. Delta-9-tetrahydrocannabinol
(delta-9-THC), endogenous cannabinoids and a multitude of
synthetic cannabinoids connect to said receptors and exert
through them an effect on the cells (Pertwee, R. G. et al.
Pharmacol. Rev. 2010, 62, 588-631). CB1 and CB2 are members of
the superfamily of the G protein coupled receptors (GPCRs). More
precisely, the receptors inhibit the adenylate cyclase via the
.. heteromeric G protein and activate the mitogenically
activated protein kinase (Howlett, A. C. et al. Pharmacol. Rev.
2002, 54, 161-202; Howlett, A. C. Handb. Exp. Pharmacol. 2005,
168, 53-79). In terms of the CB1 receptor it is further
described that it can modulate potassium flows via ion channels
of the A-type and calcium flows via N as well as P/Q-type
channels. Furthermore, CB1 receptors are able to transfer
signals to the expressing cells via Gs proteins (Glass, M.,
Felder, C. C. J. Neurosci. 1997; 17, 5327-5333; Maneuf, Y. P.,
Brotchie, J. M. J. Pharmacol. 1997; 120, 1397-1398; Calandra, B.
et al. Eur. J. Pharmacol. 1999; 374, 445-455; Jarrahian, A. et
al. J. Pharmacol. Exp. Ther. 2004, 308, 880- 886). The ability
of CB1 and CB2 to transfer signals via Gy0 and further
downstream via 2o .. inhibition of the adenylate cyclase, is
used in the so-called [35S]GTP gammaS binding assay and the cAMP
assay (Howlett, A. C. et al. Pharmacol. Rev. 2002, 54, 161-202;
Pertwee, R. G. Handb. Exp. Pharmacol. 2005a, 168, 1-51) to ana-
lyze the binding and signal transduction of cannabinoids. CB1
receptors have at their disposal an orthosteric as well as one
or multiple allosteric binding site(s), which are considered as
potential sites of action for ligands (Price, M. R. et al. Mol.
Pharmacol. 2005a, 68, 1484-1495; Adam, L. et al. 17th Annual
Symposium of the Cannabinoids, 2007, S. 86; Horswill, J. G. et
al. J. Pharmacol. 2007, 152, 805-814; Navarro, H. A. et al. J.
Pharmacol. 2009, 156, 1178-1184). CB1 receptors are mainly found
on the terminal ends of central and peripheral neurons, where
they usually impart an inhibition of excitatory and inhibitory
neurotransmitters (Howlett, A. C. et al. Pharmacol. Rev. 2002,
54, 161- CA 03023760 2018-11-09 WO 2017/194173 PCT/EP2016/060905
- 3 - 202; Pertwee, R. G., Ross, R. A. Prostaglandins Leukot
Essent Fatty Acids, 2002, 66, 101-121; Szabo, B., Schlicker, E.
Handb. Exp. Pharmacol. 2005, 168, 327-365). The distribution of
these receptors in the central nervous system is in such a way
that their activation can influence different cognitive
processes (e.g. alertness and memory, different motor functions
und pain perception). CB2 receptors are mainly localized, as
mentioned before, in immune cells. Once they get activated, they
modulate cell migration and the release of cytokines inside and
outside the brain (Howlett, A. C. et al. Pharmacol. Rev. 2002,
54, 161- 202; Cabral, G. A., Staab, A. Handb. Exp. Pharmacol.
2005, 168, 385-423; Pertwee, R. G. Handb. Exp. Pharmacol. 2005a,
168, 1-51). There is also some evidence that firstly CBI
receptors are expressed by non- neuronal cells (including immune
cells) (Howlett, A. C. et al. Pharmacol. Rev. 2002, 54, 161-202)
and that secondly CB2 receptors are expressed by some cells
inside and outside the brain (Skaper, S. D. et al. Proc. Natl.
Acad. Sci. USA 1996, 93, 3984-3989; Ross, R. A. et al.
Neuropharmacology 2001a, 40, 221-232; Van Sickle, M. D. et al.
Science 2005, 310, 329-332; Wotherspoon, G. et al. Neuroscience
2005. 135, 235-245; Beltramo, M. et al. Eur. J. Neurosci. 2006,
23, 1530-1538; Gong, J. P. et al. Brain Res. 2006, 1071, 10-23;
Baek, J. H. et al. Acta Otolaryngol 2008, 128, 961-967). Known
compounds, which have been proven to have an affinity for the
aforemen- tioned receptors CBI and CB2, are amongst others
cannabidiol (CBD) and cer- tain chemical derivatives thereof. In
particular the active compound delta-9-tetrahydrocannabinol
(delta-9-THC) from the cannabis plant has become a focus of
attention in the last couple of years. Reduced to only its
psycho-active effects in the past, recent studies show a more
diverse range of effects. Applications in cancer and HIV therapy
as well as in the treatment of multiple sclerosis are found. The
(-)-enantiomer has been found to be the more active one (Jones,
G. et al., Biochem. Pharmacol., 1974, 23: 439; Roth, S. H., Can.
J. Physiol. Pharmacol., 1978, 56: 968; Martin, B. R. et al.,
Life Sciences, 1981, 29: 565; Reichman, M. et al. Mol.
Pharmacol., 1988, 34: CA 03023760 2018-11-09 WO 2017/194173
PCT/EP2016/060905 - 4 - 823; Reichman, M. et al., Mol.
Pharmacol., 1991, 40: 547). Therefore an enantiopure product is
desirable. Since the isolation of pure trans-(-)-delta- 9-THC
from cannabis sativa or indica is a very time consuming and
expensive process (WO 2002/045115 Al), trans-(-)-delta-9-THC is
more and more synthetically produced under the name dronabinol.
This is either done in a partially synthetic way by conversion
of the precursor isolated from the cannabis plant, trans-(-)-
cannabidiol, to dronabinol (WO 2002/045115 Al) or fully
synthetic as described in EP 2842933 Bl. Different cannabinoid
compounds and methods for their manufacture are known io from
the prior art. Korte et al. (Tetrahedron Letters, 1968, 3,
145-7) describe cannabidivarin for the first time and propose a
synthesis analogous to the one by Petrzilka et al. (Hel- vetica
Chimica Acta, 1967, 50, 719-723). However, only low yields can
be achieved this way. Also Crombie et al. (Phytochemistry 1975,
4, 11975) describe the synthesis of cannabidivarin in small
scale as condensation of divarin with para- menthadienol. The
synthesis in dried CH2C12, saturated with PTSA, however, is not
very selec- tive and the resulting products are obtained at
uneconomical proportions. Tetrahydrocannabivarin (here denoted
delta-1-tetrahydrocannabivarol) is gener- ated the same way at
higher temperatures and at an uneconomical concentration in a
multiple compound mixture. WO 2006/136273 describes a method for
the manufacture of dronabinol ((denot- ed (6a R-
trans)-6a,7,8,10a-tetrahydro-6 ,6,9-trimethy1-3-penty1-6 H-
dibenzo[b,d]pyran-1-ol, g-tetrahydrocannabinol (A9-THC) in the
WO document), nowadays according to IUPAC also denoted
(6aR,10aR)-6,6,9-trimethy1-3-penty1-
6a,7,8,10a-tetrahydro-6H-benzo[c]chromen-l-ol or
delta-9-tetrahydrocannabinol, delta-9-THC or A-9-THC) from
cannabidiol (CBD) via cyclization of cannabidiol (CBD) (2-[1R-3-
methyl-6-(1-methyletheny1)-2-cyclohexene-1-y1]-5-penty1-1,3-
benzenediol) to yield delta-9-THC. The described method is
characterized in that .. cannabidiol (CBD) is provided in an
organic solvent and is heated and cyclized to delta-9-THC in the
presence of a molecular sieve. It is stated in WO 2006/136273 CA
03023760 2018-11-09 WO 2017/194173 PCT/EP2016/060905 - 5 - that
the used molecular sieve exhibits, besides the drying properties
that have been described so far, strong catalytic properties,
which are in the focus of the described conversion. Cyclizations
that can only be performed in the presence of a Lewis acid
catalyst are usually significantly slower and deliver worse
yields of delta-9-THC than cyclizations that are performed in
the presence of a molecular sieve. Further types of syntheses
are described in the literature, e.g. by Crombie et al. Chem.
Research 1977, 114, 1301-1345. More recent synthesis methods are
disclosed inter alia in EP 2314580. The method for the
manufacture of canna- binoids described therein, is supposed to
be applicable to all stereoisomers and homologs of cannabinoids
and consists of two and three chemical synthesis steps,
respectively. In a first step, alkyl resorcylic acid esters
(6-alkyl- 2,4- dihydroxybenzoic acid ester) are thereby
condensed with unsaturated hydrocar- bons, alcohols, ketones
(and their derivatives such as enol esters, enol ethers and
ketals, respectively) to the corresponding
6-alkyl-2,4-dihydroxybenzoic acid esters that are substituted at
the 3-position. In a second step, the ester function- containing
intermediates that were produced in the first step are subjected
to a decarboxylating saponification, giving rise to the
corresponding ester-free canna- binoids. If necessary, an acid
catalyzed rearrangement is carried out in a third step. This
isomerization may be e.g. the ring closure of the pyran ring of
CBD to give dronabinol, but also the rearrangement of a double
bond like e.g. the reor- ganization of delta-9 to delta-8-THC or
an acid catalyzed epimerization like the rearrangement of
cis-9-ketocannabinoids to the corresponding trans-compounds. US
5,342,971 describes a method for the manufacture of dronabinol
and of the related dibenzo[b,d]pyrans. These are produced,
according to the abstract, through heating of a dihydroxybenzoic
acid derivative in the presence of a Lewis acid catalyst and an
inert non-polar solvent, in which indeed the dihydroxybenzoic
acid is soluble, but the Lewis acid catalyst is insoluble or
only very slightly soluble. .. EP 2842933 B1 discloses a method
for synthesizing delta-9-THC starting from menthadienol. In a
first step, menthadienol is reacted with an olivetolic acid
ester to a cannabidiolic acid ester. This ester is then
subjected to a transesterification CA 03023760 2018-11-09 WO
2017/194173 PCT/EP2016/060905 |EPO DP num="6"|
[0003] and the product is saponified and decarboxylated to
cannabidiol. In the last step, cannabidiol is cyclised to
trans-(-)-delta-9-THC in enantiopure form. oH a) EiF3*()Etz
TBME, RT. 3t1 OH > b) chromatographic purification = /HO 60%
(I) (II) (III) menthadienol cannabidiol
trans-(-)-delta-9-tetrahydrocannabinol Details of the synthesis
of delta-9-THC according to EP 2842933 B1 can be found in
example 1. Analogously, a synthesis of cannabidivarin (CBDV) and
tetrahydrocannabivarin (THCV) starting with reacting
menthadienol with a divarinic acid ester, followed io by
transesterification, saponification and decarboxylation to
cannabidivarin and subsequent cyclisation to
tetrahydrocannabivarin is described in European patent
application EP 15156750Ø The product is also obtained in
enantiopure form as trans-(-)-delta-9-THCV. An example of the
synthesis steps for cannabidivarin (CBDV) and
tetrahydrocannabivarin (THCV) as described in European patent
application EP 15156750.0 is shown schematically below. Reaction
conditions can be in- ferred from example 2. CA 03023760
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[0004] 1. Coupling step: OH OH 0 itV 111 + OH 0 1 010 HO =>i
HO (I) (IV) (V) menthadienol divarinic acid methylester ..
cannabidivarinic acid methylester 2. Transesterification step:
OH 0 OH 0 % HO % HO (V) (VI) cannabidivarinic acid methylester
2-hydroxyethylcannabidivarinolate 3.
SaDonification/decarboxylation step =H 0 OH 140 -a. HO ./ .1 HO
===>1 (VI) (VII) 2-hydroxyethylcannabidivarinolate
cannabidivarin CA 03023760 2018-11-09 WO 2017/194173
PCT/EP2016/060905 |EPO DP num="8"|
[0005] 4. Cyclization step 401 'H 1110 CH 110 7HO -J**ttl 1101
(VII) cannabidivarin trans-(-)-delta-9-tetrahydrocannabivarin
The raw product generated by the above mentioned synthesis
according to EP 2842933 B1 has a delta-9-THC content of 65 - 75
%, as well as 20 - 30 % of the isomer
delta-8-tetrahydrocannabinol as main impurity. The purification
of trans-(-)-delta-9-THC proves difficult because it can not be
obtained in crystalline form. Pure delta-9-THC is a slightly
yellow, air- sensitive resin. Therefore, a crystallization as
with the related cannabinoids, cannabidiol or cannabidivarin is
not possible when an enantiopure product is desired.
Distillation is also not feasible due to the high boiling point
(about 200 C at 0.02 mbar) and its thermal instability. A
particular complication is the presence of structurally very
similar compounds with almost identical chemical and physical
properties (polari- .. ty, boiling point etc.), which may impede
purification, such as the two isomers
delta-8-tetrahydrocannabinol and
delta-9(11)-tetrahydrocannabinol. H OH A8 - Tetrahydrocannabinol
A9(11)- Tetrahydrocannabinol The same problems arise for the raw
product obtained by the synthesis of THCV as described above,
where the corresponding isomers are formed. CA 03023760
2018-11-09 WO 2017/194173 PCT/EP2016/060905 |EPO DP num="9"|
[0006] For these reasons, expensive and time consuming
chromatographic methods are employed. WO 2002/062782 Al
discloses a method for the production of dronabinol starting
with the isolation of cannabidiol from fibrous hemp, which is
then chemically cyclised. According to the examples of WO
2002/062782, the resulting reaction mixture comprises up to 86 %
of dronabinol, which is then isolated chromato- graphically on a
silica gel column. The solvent is removed and the product is
purified by high vacuum distillation or crystallization.
However, as indicated above, distillation of dronabinol is
highly inefficient because of its thermal instabil- ity and
crystallization is only possible with an enantiomeric mixture.
According to WO 2009/133376 Al, delta-9-THC and delta-9-THC
carboxylic acid are extracted from plant material and then the
delta-9-THC carboxylic acid is converted to delta-9-THC in the
same solvent. For further purification, the product is run over
a charcoal column, the fractions containing delta-9-THC are com-
bined, concentrated and then purified by reverse phase
chromatography. Again, the combined fractions containing the
product are concentrated, extracted with MTBE and filtered.
Ethanol is added to the filtrate and the solution is concentrat-
ed to produce an oil, from which the solvent is evaporated. US
2015/0126596 Al relates to methods for producing
trans-(-)-delta-9-THC and trans-(+)-delta-9-THC in several
different ways. In one case, the preparation is started with an
enantiomeric mixture, where the two enantiomers are purified
together by preparative HPLC and then crystallized as a mixture
after which, in a resolving step, the (+/-)-enantiomers are
separated by chiral chromatography. Another way starts also with
an enantiomeric mixture, which is reacted to a nitro- benzene
sulfonate, crystallized and reacted back to a clean enantiomeric
mixture, which is then again separated by chiral chromatography.
A further way is de- scribed, in which the two separate
enantiomers are synthesized, mixed for crys- tallization and
subsequently separated again by chiral chromatography. To purify
trans-(-)-delta-9-THC by crystallization with the (+)-enantiomer
and subsequent chiral separation, however, appears to be a very
complicated purification method, which is bound to result in a
considerable loss of material and hard to scale up in an
efficient way. CA 03023760 2018-11-09 WO 2017/194173
PCT/EP2016/060905 |EPO DP num="10"|
[0007] Preparative HPLC is only successful on a very small scale
when large losses of yield are to be avoided. Only small amounts
of raw product (25 mg) can be sepa- rated into Dronabinol and
the main impurity, delta-8-THC, as demonstrated in Figures la)
and 1b). Fig. la) shows the preparative HPLC purification of 25
mg raw product obtained in the synthesis according to EP 2842933
B1 , wherein the two peaks are dronabinol as main product
(larger peak) and delta-8-THC as main impurity (smaller peak).
Fig. 1 b) shows the preparative HPLC purification of 200 mg raw
product comprising dronabinol as main product and delta-8-THC as
main impurity, demonstrating that these compounds can not be
resolved in this quanti- ty. A further chromatographic
purification method is super critical fluid chromatog- raphy
(SFC), in which liquid CO2 is used as eluant. The purification
of (-)- delta-9- trans-THC by SFC is described in WO 2005/061480
Al. This process, however, requires a complex constructional
setup and is very expensive. Moreover, the CO2 evaporates and is
lost during processing. Further methods are derivatisations of
dronabinol or precursors thereof to com- pounds which may be
crystallized. In order to perform a crystallization, the raw
product is converted into a crystallizable derivate, which is
then purified by crys- tallization and finally converted back to
dronabinol in a chemical conversion step. The most relevant
methods comprise the derivatisation of delta-9-THC to suitable
crystallizable salts, subsequent crystallization and thermal
decarboxylation to dronabinol as described in WO 2013/045115 Al.
A further option is the derivatisation of raw dronabinol to a
1-naphtoyl ester, subsequent crystallization and finally
saponification to pure dronabinol (see WO 2006/007734 Al). In
summary, the methods to purify cannabinoid compounds available
in the prior art are fairly complicated, time-consuming and
expensive. Moreover, certain impurities derived e.g. from
synthetic preparation steps, which may be structurally very
similar such as isomers of the desired product, can not be
removed to a satisfactory degree. This is especially true when
the process is to be carried out on an economically relevant
scale. CA 03023760 2018-11-09 WO 2017/194173 PCT/EP2016/060905
|EPO DP num="11"|
[0008] As a result, there is still a need to provide a method
for the purification of canna- binoid compounds, which is
suitable to achieve a maximum degree of purity while at the same
time allowing purification on a large scale in an economically
appro- priate, i.e. a time and cost efficient way. Simulated
moving bed chromatography (SMB chromatography) is a continuous
process based on the true moving bed principle, in which the
solid phase moves in the opposite direction to the liquid phase
and is therefore not stationary. Due to this opposing movement
two pure compounds can be isolated or a pure com- pound can be
isolated from a complex mixture. io The moving solid phase on
which this concept relies, however, is technically not feasible
and therefore simulated. This is implemented by arranging
several pre- parative columns connected in series and
periodically changing the valve setting so that a movement of
the solid phase in the opposite direction of the flow of the
liquid phase is simulated. The system is continuously fed with a
feed mixture comprising the compounds to be separated and an
eluant while a raffinate and an extract are continuously
withdrawn from the system. The system is therefore divided into
four different separation zones, in each of which the same
number of columns are distributed. The process shown in Figure 2
comprises 8 columns in total, but alternatively, only four may
be used. By periodically switching the feed, eluant, extract and
raffinate ports in the same direction, each column passes
through each zone once per cycle. The feed mixture is fed into
the system between zones II and III, in which the actual
separation occurs. Zones I and IV are regeneration zones. The
parameters, which are important for the SMB principle, are the
periodical change of the position of the ports as well as the
different flow rates in the four zones. These four flow rates
are regulated by four pumps. The extract pump in zone II and the
raffinate pump in zone IV are inside the column circle, the
eluant and the feed pump are located outside of the column
circle. A fine regulation is achieved by two needle valves,
which regulate the ratio between the circle flow and the outlet
flow. CA 03023760 2018-11-09 WO 2017/194173 PCT/EP2016/060905
|EPO DP num="12"|
[0009] While the prior art does not provide a process for the
purification of cannabinoid compounds, which is suitable to
achieve a purity level comparable to the process according to
the present invention as described below, in particular when
per- formed at a preparative scale, it has now been found out,
that the purification of cannabinoid compounds using a simulated
moving bed chromatographic system, preferably in combination
with one or more additional extraction step(s), provides one or
two desired cannabinoid products with an unexpectedly high
degree of purity while still allowing the process to be
implemented on an economically relevant scale. io This finding
was unexpected as conventional silica gel chromatography of
larger amounts of dronabinol fails to provide a viable option to
separate the product from its isomers, while it is on the other
hand not feasible to scale up reversed phase HPLC chromatography
to preparative significant amounts. The method according to the
invention, however, surprisingly compensates both theses prob-
lems and provides a way to obtain pure product on a large scale.
It was therefore an objective of the present invention to
provide a purification method which overcomes the above
mentioned problems. In particular, it was an objective of the
present invention to provide a purification process for one or
two cannabinoid compound(s) from a reaction mixture derived 2o
from a synthetic preparation process, especially a process as
described in EP 2842933 B1 . It was also an object of the
present invention to obtain the desired cannabinoid compound(s)
in a degree of purity that any of its/their isomers are below a
detec- tion level and furthermore in enantiopure form. The
objectives given above are met by a method for purifying one or
two can- nabinoid compounds comprising the steps: i) providing a
mixture comprising at least one cannabinoid compound obtained by
enantiopure synthesis and one or more of its isomers and
optionally one or more further organic compounds, and CA
03023760 2018-11-09 WO 2017/194173 PCT/EP2016/060905 |EPO DP
num="13"|
[0010] ii) simultaneously, a) continuously feeding the mixture
of step i) through a feed port into a simulated moving bed
chromatographic apparatus com- prising at least four columns
connected in series and containing a stationary phase, and b)
continuously feeding eluant into the apparatus through an eluant
port, and c) continuously withdrawing the extract through an
extract port, and d) continuously withdrawing the raffinate
through a raffinate port, wherein the extract and/or the
raffinate respectively comprise(s) one purified cannabinoid
compound and wherein the extract and/or the raffinate comprising
one purified cannabinoid compound comprise(s) less than 100 ppm,
preferably less than 70 ppm, particularly preferably less than
50 ppm in total of any iso- mer(s) of the purified cannabinoid
compound present in step i). As described above, SMB
chromatography allows separation and purification of one or
simultaneously two desired product compounds, the stronger
adsorbing compound is obtained as the extract and the weaker
adsorbing compound is obtained as the raffinate. Advantageously,
a mixture comprising at least one cannabinoid compound together
with at least one of its isomers, such as a reac- tion mixture
from a synthesis step, may be subjected to the method according
to the invention to provide highly pure products in large
yields. The at least one cannabinoid compound and its isomer(s)
present in the mixture provided in step i) are likely very
similar in chemical structure and therefore also in their
physical properties. Consequently, they are particularly hard to
separate. Using the meth- od according to the present invention,
however, the cannabinoid compound(s) can efficiently be
separated from their isomers. CA 03023760 2018-11-09 WO
2017/194173 PCT/EP2016/060905 |EPO DP num="14"|
[0011] The one or more further organic compound(s) present in
the mixture provided in step i) may be any compound(s) selected
from synthetic starting materials or side products of the
synthesis, which are not cannabinoid compounds. As the steps a)
to d) are carried out simultaneously and continuously, the
process is very time efficient and the required amount of eluant
is significantly reduced compared to conventional
chromatography. Additionally, it is advantageous that the solid
phase can be used for separation during the entire process. This
in- creases the efficiency of the separation and simultaneously
reduces the required amount of eluant. Once the adsorption
equilibrium is reached, the composition of io .. the raffinate
and extract do not change anymore as long as the respective pa-
rameters are not changed. The loss of valuable materials is
reduced to <5%. As a simulated moving bed chromatographic
apparatus, any system suitable to perform simulated moving bed
chromatography may be used. The stationary or solid phase may be
any material, which the skilled person can easily choose
according to the nature of the mixture and the compounds to be
separated. To determine the respective flow rates at the
different pumps, several methods and models are known in the
prior art which may be implemented in order to achieve an
optimal separation of the desired compound(s). The desired
compound(s) is/are obtained in the extract and/or the raffinate
in a degree of purity with respect to any isomer of the desired
cannabinoid com- pound(s), which is preferably below detection
limit, in particular the total amount of any isomer of the
cannabinoid compound(s) which was/were present in step i) is
less than 100 ppm, preferably less than 70 ppm and particularly
preferably less than 50 ppm. The degree of purity may be
determined chromatographically using an HPLC apparatus (e.g.
Knauer HPLC smartline series) and the appropriate USP reference
standards for dronabinol and its isomers. A Restek - Raptor (ARC
- 18, 2.7 mm, 150 x 4.6 mm) HPLC column may be used together
with the re- spective USP eluent (45 % methanol, 25 % water, 20
% tetrahydrofuran, 10 % acetonitrile) with an eluent flow of 0.8
mL / min. .. According to a further aspect the method as
described above additionally com- prises the step CA 03023760
2018-11-09 WO 2017/194173 PCT/EP2016/060905 |EPO DP num="15"|
[0012] iii) subjecting the extract and/or the raffinate
comprising one purified cannabinoid compound to one, two or more
further extraction step(s), preferably using an oil as the
extracting agent, wherein the extract and/or the raffinate
respectively obtained in step iii) com- prise(s) one purified
cannabinoid compound and less than 100 ppm, preferably less than
70 ppm, particularly preferably less than 50 ppm in total of any
further organic compound(s) present in step i). By subjecting
the desired cannabinoid compound(s) which is/are contained in
the extract and/or the raffinate to one or more further
extraction step(s), impurities io other than the isomers present
in step i) can be removed. In particular, organic compounds,
which may be present from the synthetic step such as starting
mate- rials or side products of the synthesis can be removed to
a degree such that they are present in an total amount of less
than 100 ppm, preferably less than 70 ppm, particularly
preferably less than 50 ppm. The extraction agent may be any
substance selected from the group consisting of cyclohexane,
heptane and other oxygen free hydrocarbons. Suitable oils to be
used as extracting agents are selected from the group consist-
ing of plant oils with medium chain triglycerides, preferably
containing the fatty acids capric acid and caprylic acid.
Advantageously, when using an oil as extract- ing agent, the
resulting product comprising the desired cannabinoid compound ¨
besides being highly pure ¨ is particularly stable when kept
under argon and in the dark. In particular, the medium chain
triglycerides mentioned above provide an antioxidant effect and
therefore enhance the stability of the product. In the method
according to the invention, the cannabinoid compound(s) to be
purified may be selected from the group consisting of
cannabidiol, trans-(-)- delta- 9-tetrahydrocannabinol,
cannabidivarin, trans-(-)-delta-9- tetrahydrocannabivarin and
cannabigerol. Any of these compounds may be obtained by chemical
synthesis as described in the prior art, which results in
reaction mixtures comprising one or more canna- binoid compounds
such as the desired product and its synthetic precursors,
as
[0013] well as at least one isomer of the product compound(s)
and further organic com- pounds such as starting materials or
side products of the synthesis, which are not cannabinoid
compounds. These mixtures can be purified to a high degree and
with good yields on a large scale by the method according to the
present inven- tion. In a particularly preferred embodiment, the
method according to the invention is used to purify the reaction
product(s) of the synthesis steps described in EP 2842933 B1 .
The reaction product(s) to be purified are preferably trans-(-)-
delta- 9-THC (III) or cannabidiol (II). io According to one
aspect, in the method according to the present invention step i)
includes the following step: conversion of menthadienol with an
olivetolic acid ester to a cannabidiolic ac- id ester of formula
(IX) OHO 0 'IFIO (IX) wherein Y is an organic residue,
preferably in a continuous process. According to a further
aspect, step i) comprises the conversion of a cannabidiolic acid
ester of formula (IX), wherein Y is an organic residue with an
alcohol of the formula HO-X, wherein
[0014] X is an aliphatic residue with one, two, three or more
than three hydroxyl groups, wherein the total number of C-atoms
in the aliphatic residue X is not greater than 15, and wherein
the aliphatic residue is - saturated or unsaturated and -
branched or unbranched, wherein Y is different from X and
selected such that the alcohol of formula HO- Y, which is
generated during the conversion, boils at a lower temperature at
1013 hPa than the used alcohol of formula HO-X. According to yet
another aspect of the method according to the invention, the
compound generated by the conversion of the cannabidiolic acid
ester of formula (IX) with the alcohol of formula HO-X is
treated in such a way that it is decarboxylated and saponified
to generate cannabidiol (II). In a further aspect of the method
according to the invention, the cannabidiol, which is present
after the decarboxylating saponification, is cyclised to
trans-(-)- delta-9-tetrahydrocannabinol (III), preferably in the
absence of halogenated sol- vents. In the synthesis of
delta-9-tetrahydrocannabinol according to EP 2842933 B1, an 2o
impurity which is very hard to remove is olivetol, which is
generated during the synthesis. Surprisingly, when the method
according to the present invention is used to purify the product
delta-9-THC in combination with the one or more further
extraction step(s), any residual olivetol can be removed to such
a degree that it is no more detectable by conventional HPLC
analysis as demonstrated in example 3.
[0015] Therefore the present invention particularly relates to a
method as described above, wherein the one or one of the further
organic compound(s) present in step i) is olivetol. Furthermore,
as already mentioned in the introduction, the raw product
generated by the synthesis according to EP 2842933 B1 has a
delta-9-THC content of 65 ¨ 75 %, as well as 20 ¨ 30 A of the
isomer delta-8-tetrahydrocannabinol as main impurity.
Furthermore, delta-9(11)-tetrahydrocannabinol may be present.
Due to the structural similarity of the isomers, the desired
delta-9-THC is very hard to purify from this reaction mixture.
io Using the method according to the present invention, however,
a purity may be obtained such that these isomers can no more be
detected in the product as demonstrated in example 3. Therefore
in a particularly preferred embodiment, in the method according
to the invention, the mixture provided in step i) comprises
trans-(-)-delta-9- tetrahydrocannabinol together with
delta-8-tetrahydrocannabinol and/or delta- 9(11
)-tetrahydrocannabinol. According to a further preferred
embodiment, the method according to the inven- tion is used to
purify the reaction product(s) of the synthesis steps described
in European patent application EP 15156750Ø The reaction
product(s) to be pun- 2o fied are preferably cannabidivarin
(VII) or trans-(-)-delta-9- tetrahydrocannabivarin (VIII).
According to one aspect in the method according to the present
invention, step i) comprises the conversion of menthadienol of
formula (I) with a divarinic acid ester of formula (IV), to an
ester of formula (V),
[0016] OH OH 0 OH 0 001 + HO HO 14111 (I) (IV) (V) According to
a further aspect, in the method according to the invention, step
i) comprises the transesterification of the ester of formula (V)
with an alcohol of the formula HO-X, wherein X is an aliphatic
residue with no, one, two, three or more than three hy- droxyl
groups, wherein the total number of C-atoms in the aliphatic
resi- due X is not greater than 15, and io wherein the aliphatic
residue is - saturated or unsaturated and - branched or
unbranched, - acyclic or cyclic, with the proviso that the
alcohol of formula HO-X is selected from the group consisting of
cyclohexanol and hexanol in case X is an aliphatic residue with
no hydroxyl group. According to yet a further aspect, in the
method according to the invention, the compound generated by the
conversion of ester of formula (V) with the alcohol of
[0017] formula HO-X is treated in such a way that it is
decarboxylated and saponified to generate cannabidivarin (VII).
In a further aspect of the method according to the invention,
the cannabidivarin, which is present after the decarboxylating
saponification, is cyclised to trans-(-)-
delta-9-tetrahydrocannabivarin (VIII), preferably in the absence
of halogenated solvents. The reaction products of the synthesis
steps as described in European patent application EP 15156750.0,
may be purified by the method according to the invention. The
advantages described above in the context of the purification of
io the synthesis product(s) according to EP 2842933 apply
accordingly. Finally, the present invention also relates to an
extract or raffinate obtained or obtainable in step c) or d) of
a method as described above, or an extract or raffi- nate
obtained or obtainable in step iii) of a method as described in
the context of the corresponding embodiment above. An extract or
raffinate obtained or obtainable in step c) or d) of a method as
described above, or an extract or raffinate obtained or
obtainable in step iii) of a method as described in the context
of the corresponding embodiment above, comprises the desired
cannabinoid compound in a degree of purity, in particular with
respect to its isomers, which could not be achieved by any of
the conven- tional processes available in the prior art. The
following examples describe particular embodiments of the
present inven- tion, without meaning to limit the scope of
protection. Example 1: Synthesis of delta-9-THC: Step 1:
Coupling step (in the continuous process); Synthesis of
cannabidiolic acid methyl ester (I)
[0018] a)13F3=0E12, H chi ro b enzene, PT b) NaHCO3, S --- PT,
20 min OH a o 40 - 70% Iv HO 'girl' a H menthadienol olivetolic
acid cannabidiolic acid methyl ester methyl ester 300 g (2.0
mol) menthadienol and 476 g (2.0 mol) olivetolic acid ester are
dis- solved at ca. 22 C in 1,370 g of chlorobenzene (2,000 mL
solution A), likewise 94 g (0.66 mol) boron trifluoride*etherate
are dissolved in 640 g of chlorobenzene at ca. 22 C (666 mL
solution B)., Solution A at a flow rate of 72 mU min and solu-
tion B at a flow rate of 24 mL/ min are pumped into a stirred
reaction chamber via two separate dosing pumps, from the
reaction chamber the reaction composition runs via a PTFE hose
into a stirred solution of 1,000 g of sodium bicarbonate. The
total reaction time is ca. 20 min. After termination of the
metering the hydro- lyzed reaction solution is stirred for a
further 30 min. Then the hydrolyzed reaction solution is
transferred into a 5L jacket reaction vessel, the aqueous phase
is separated and the solvent chlorobenzene is re- moved in
vacuo. Ca. 2,000 g of toluene are added to the remaining 730 g
of raw material and the unreacted olivetolic acid ester is
extracted through the addition of 1,200 g 1% aqueous sodium
hydroxide solution (four times). After acidifying with semi
conc. sulfuric acid and re-extraction of this aqueous phase, ca.
30% (140 g) of non converted olivetolic acid ester are
recovered. There are ca. 520 g of cannabidiolic acid methyl
ester in the toluene phase, which corresponds to a theoretical
yield of ca. 70%. This first intermediate serves as starting
material for the following transesterification. Step 2:
Transesterification, synthesis of 2-hydroxyethyl
cannabidiolate:
[0019] /00 C*-1 = KOH, ethylene ghicol OH 0.5 bar,120 0, 2h z
/HO --7H0 2-hydra xyethyl cannabidiolate The toluene is removed
in vacuo and to the remaining first intermediate 600 g of
ethylene glycol are added under stirring followed by a solution
of 85 g of potassi- um hydroxide in 300 g ethylene glycol. A
vacuum of ca. 0.5 bar is applied and it is heated to 120 C for 2
h, whereby ca. 40 g of methanol distill off. The resulting
product composition mainly comprises 2-hydroxyethyl
cannabidiolate. Step 3: Saponification/ decarboxylation,
synthesis of cannabidiol (X): OH OH 0.5 bar, 150*C, 211 0 60%
cannabldol Subsequently, the temperature is increased to 150 C
and it is stirred at this io temperature for 2 h. The product
composition resulting from the transesterification comprising
mainly 2-hydroxyethyl cannabidiolate is cooled down to ca. 40 C
and 500 g of water as well as 500 g of n-heptane are added and
ca. 150 g of semi conc. sulfuric acid are added for
neutralization. After phase separation, the solvent is removed
using a rotary evaporator and the remainder is distilled over a
thin-film evaporator using a vacuum of ca. 0.5 mbar and a jacket
temperature of 230 C. 310 g of cannabidiol are obtained in the
form of a viscous, yellowish oil with a purity of 85%, which
corresponds to a theoretical yield of 60% in relation to the
used cannabidiolic acid ester.
[0020] This viscous, yellowish oil is then recrystallized in ca.
200 g of n-heptane at ca. -5 C, after which 210 g of white
crystallizate with a purity of 99% cannabidiol are obtained.
Step 4: Cyclization. synthesis of delta-9-THC: SI a) BF3*OEtz,
TBME, RI, 3h OH b) chromatographic Purification /HO a 60%
cannabidiol d eita-9-THC 50 g of pure cannabidiol are dissolved
in 250 g methyl-tert-butylether and 40 g of boron
trifluoride*acetic acid complex are added under stirring within
10 min at ca. 22 C. It is stirred for 3 h at said temperature
and then 200 g of ice water are added, the organic phase is
washed with sodium bicarbonate solution and the io solvent is
removed using a rotary evaporator. The remaining raw material of
ca. 50 g contains 74% trans-(-)-delta-9-tetrahydrocannabinol
(delta-9-THC), 25% of side products as well as < 1%
cannabidiol. Example 2: Synthesis of cannabidivarin and
tetrahydrocannabivarin: Step 1: Coupling step OH OH 0 + (3* = H
= HO 101 HO (I) (IV) (IV)
[0021] 273g (1,8 Mol) menthadienol and 377g (1,8 Mol) divarinic
acid methylester are dissolved at RT in 1.450g toluene (2.300mL
solution A), likewise, an adequate amount of
borontrifluoride*etherate are dissolved in 540g toluene at RT
(710mL solution B). Solution A and solution B are pumped into a
stirred reaction chamber via two separate dosing pumps, from the
reaction chamber the reaction composi- tion runs via a PTFE hose
into a stirred solution of 1,000 g of sodium bicar- bonate. The
total reaction time is about 25 mins. After termination of the
metering the hydrolyzed reaction solution is stirred for about 1
hour. Then the hydrolyzed reaction solution is transferred into
a 5L jacket reaction vessel, the aqueous phase is separated. The
not reacted divarinic acid ester is extracted by six times
adding 1.000g of 1% aqueous sodium hydroxide solution. After
acidifying with semi conc. sulfuric acid and re-extraction of
this aqueous phase, ca. 30% (130 g) of non converted divarinic
acid ester are recovered. In the toluene phase, about 320g
cannabidivarinic acid methylester (V) are con- tamed, which
corresponds to a theoretical yield of 50%. This first
intermediate serves as starting material for the following
transesterification. Step 2: Transesterification step: OH 0 OH 0
OH : .......I (V) (VI) 2o The toluene is removed in vacuo and to
the remaining first intermediate 650 g of ethylene glycol are
added under stirring followed by a solution of 122 g of potas-
sium hydroxide in 420 g ethylene glycol. A vacuum of ca. 0.5 bar
is applied and it is heated to 100-120 C for 2 h, whereby ca. 40
g of methanol distill off. The resulting product composition
mainly comprises 2-hyd roxy- ethyl- cannabidivarinolat (VI).
[0022] Step 3: Saponification / Decarboxylation: OH 0 OH LUOH
41111 ..!7* HO >4 HO (VI) (VII) Subsequently, the temperature
is increased to 150 C and it is stirred at this temperature for
3-4h (also in vacuo; cfl. step 2). The product composition
result- ing from the transesterification is cooled down to ca.
40 C and 1.500 g of water as well as 800 g of methyl-tert.
butyether are added and ca. 180 g of semi conc. sulfuric acid
are added for neutralization. After phase separation, the
solvent is removed using a rotary evaporator and the remainder
is distilled over a thin- film io .. evaporator using a vacuum
of ca. 1 mbar and a jacket temperature of 230 C. 270 g of
cannabidivarin (VII) are obtained in the form of a viscous,
yellowish oil with a purity of 85%, which corresponds to a
theoretical yield of 85% in relation to the used cannabivarinic
acid ester. This viscous, yellowish oil is then recrystallized
in ca. 270 g of n-heptane at ca. 10 C, after which 190 g of
white to lightly yellow crystallizate with a purity of 99%
cannabidivarin (VII) are obtained. Step 4: Cyclization to
tetrahydrocannabivarin (THCV): 50 g of pure cannabidivarin (VII)
are dissolved in 250 g methylene chloride and 40 g of boron
trifluoride*ether complex are added under stirring within 10 min
at ca. 22 C. It is stirred for 20 mins at said temperature and
then 200 g of ice water are added, the organic phase is washed
with sodium bicarbonate solution and the solvent is removed
using a rotary evaporator. The remaining raw material of ca. 50
g contains 74% trans-(-)-delta-9-tetrahydrocannabivarin (VIII)
and 26% of side products.
[0023] Example 3: Purification of a raw product as obtained in
example 1: Any steps described herein were conducted in an inert
gas atmosphere (argon) due to the air-sensitivity of the
dronabinol. After processing the reaction mixture, the following
composition of the raw product is obtained: HPLC-Analysis: (DAD,
in Area-%) substance ¨ batch number LN 703795 LN 703814 LN
703842 olivetol (2,8 min) 1,2% 1,3% 1,2% cannabidiol (8,5 min)
0,3% 0,4 % 0,4 % dronabinol (14,8 min) 71,6 % 71,4 % 72,1 % _
6,9(11)-tetrahydrocannabinol (15,6 min) 0,4 % 0,4 % 0,4 % _
6,8-tetrahydrocannabinol (17,0 min) 26,3 % 26,3 % 25,4 % _
Figure 3 shows the exemplary chromatogram of LN 703795. The
chromatographic system is based on a known SMB apparatus of the
com- pany Knauer (Germany). The system comprises 8 separation
columns (Knauer Vertex Plus, 250 x 8 mm), as well as the
required pumps. The column configura- tion corresponds to the
standard 2 ¨ 2 ¨ 2 ¨ 2 arrangement. The movement of the
individual columns is implemented by a 64 port rotary valve. The
switching time of the valve is 10.81 seconds. The valve and the
HPLC columns are located in a tempered column oven. The
temperature of the chromatographic system is to 60 C, preferably
30 C. 15 A solid phase suitable for the separation is an RP
material (Eurospher ll silica gel, C18P) with a grain size of 10
to 100 pm, preferably 20 to 45 pm. The solid phase showed no
signs of deterioration over a time period of two years. This is
a further advantage compared to classical chromatographic
systems. As mobile (liquid) phase / eluant a mixture of
methanol, tetrahydrofuran and 20 water is used, preferably with
the composition: methanol (62%), tetrahydofuran
[0024] (17%), water (21%). Furthermore, 0.01% ascorbic acid is
added to the mixture as antioxidant. The feed mixture comprises
the above described raw product dissolved in eluant mixture at a
concentration of 12.5 g/L. The eluant, extract and raffinate
pump .. each have a maximum flow rate of 50 ml/min, the feed
pump a maximum flow rate of 10 ml/min. In the process described
herein, the following flow rates are used: eluant pump (zone 1;
4.4 ml/min), extract pump (zone 2, 3.2 ml/min), raffi- nate pump
(zone 4, 1.3 ml/min) and feed pump (zone 3, 0.2 ml/min). The
flow rates are measured with a Humonics Optiflow 520. io The
supply of the system with eluant and feed solution is done from
suitable stock containers, which are secured for fire safety.
Eluant and feed solution are periodically overlaid with argon to
keep oxygen from the air out. Before entering the system, eluant
and feed solution are pumped though a deaerator. The SMB process
does not need a constant supervision. The process described
herein may be run continuously over several weeks without
changing the pa- rameters and without having a change in the
yield. The particular stability of the process allows a 24 hour
operation, without needing shift workers. Internal con- trols of
the process are performed once a day. With the process described
herein, 0.15 g/ hour of dronabinol can continuously be obtained
from the raffinate. This corresponds to a daily rate of 3.6 g.
By upscaling the process from 8 mm to 50 mm columns, the daily
yield can be increased to 144 g of pure dronabinol. This
corresponds to a yearly production of about 40 kg of dronabinol.
The raffinate derived from the SMB process has the following
composition: HPLC-Analysis: raffinate (DAD, in Area-%)
[0025] substance ¨ batch number LN 703795 LN 703814 LN 703842
olivetol 1,1% 1,1% 1,4% cannabidiol 0,4 % 0,2 % 0,4 % dronabinol
>97 % >97 % >97 % 6,9(11) - tetrahydrocannabinol n.d.
n.d. n.d. 6,8 - tetrahydrocannabinol n.d. n.d. n.d. n.d. = not
detectable Figure 4 shows the exemplary chromatogram of the
raffinate from LN 703795. After adjusting the adsorption
equilibrium, the obtained raffinate is subjected to further
processing. The solvent is reduced by distillation (100 mbar
vacuum at a temperature of 30 C) to 30 % organic. The distilled
solvent is reintroduced to the process as eluant after
adjustment of the starting mixture. The obtained reduced
raffinate is extracted twice with cyclohexane (50 wt.-% with
respect to the re- duced raffinate). The olivetol contained in
the raffinate stays in the water/organic phase, while the
dronabinol passes into the cyclohexane phase. After removal of
the solvent by distillation, dronabinol is obtained with a
content of > 99% at a residual solvent content of below 100
ppm. HPLC-Analysis: final product (DAD, in Area-%) substance ¨
batch number LN 703795 LN 703814 LN 703842 olivetol n.d. n.d.
n.d. cannabidiol 0.31 % 0.51 % 0.41 % dronabinol 99.34 % 98,10 %
99.15 % 6,9(11) - tetrahydrocannabinol n.d. n.d. n.d. 6,8 -
tetrahydrocannabinol n.d. n.d. n.d. n.d. = not detectable
[0026] Figure 5 shows the exemplary chromatogram of the final
product from LN 703795. For extraction, instead of cyclohexane,
a plant oil based on a mixture of medium chain triglyceride may
alternatively be used. This leads to a comparable purity as
obtained with cyclohexane and a stable storage medium for the
pure compound. Short description of the drawings: Fig. la) shows
the preparative HPLC purification of 25 mg raw product obtained
in the synthesis according to EP 2842933 B1, wherein the two
peaks are dronabinol as main product (larger peak) and
delta-8-THC as main impurity io (smaller peak). Fig. 1 b) shows
the preparative HPLC purification of 200 mg raw product obtained
in the synthesis according to EP 2842933 B1 comprising
dronabinol as main product and delta-8-THC as main impurity,
which can not be resolved in this quantity. Figs. 2a) and b)
show a schematic setup of a SMB system. Figure 3 shows the
exemplary chromatogram of LN 703795. Figure 4 shows the
exemplary chromatogram of the raffinate from LN 703795. Figure 5
shows the exemplary chromatogram of the final product from LN
703795.
Cannabidiol
synthesis method
CN106810426
The invention provides
a cannabidiol synthesis method. The method includes: taking
2,4-dyhydroxy-6-pentyl methyl benzoate as a raw material, and
subjecting to ester exchange with N,N-dialkyl alcohol amine
under potassium hydroxide catalysis; subjecting to coupled
reaction with (1S,
4R)-1-methyl-4-(1-methylvinyl)-2-cyclohexene-1-alcohol under
Lewis acid catalysis; performing acid-base extraction and
recrystallization to obtain a high-purity key intermediate
product; subjecting to hydrolysis decarboxylation to obtain
crude cannabidiol, and subjecting the crude cannabidiol to once
recrystallization to obtain cannabidiol according with raw
medicine quality requirements. The cannabidiol synthesis method
has advantages that raw materials and reagents are cheap and
commercially easy to acquire, the total yield of the finally
prepared raw medicine in qualified purity reaches 35-40%, the
process is improved evidently, and the method has a promising
industrial application prospect.
The present invention provides a transesterification of
N,N-dialkylolamine catalyzed by potassium hydroxide with
2,4-dihydroxy-6-pentyl benzoic acid methyl ester as a raw
material, and then The coupling reaction of
1S,4R)-1-methyl-4-(1-methylvinyl)-2-cyclohexene-1-ol is
catalyzed by Lewis acid, and is obtained by acid-base extraction
and recrystallization. The key intermediate product of purity is
hydrolyzed and decarboxylated to obtain crude cannabisdiol. The
crude product can be recrystallized at one time to obtain
cannabisdiol which meets the quality requirements of the drug
substance. The raw materials and reagents in the method of the
invention are low in price and commercially available, and the
total yield of the qualified raw materials obtained in the final
method is up to 35-40%, and the process is obviously improved,
which has a good industrial application prospect.
Background
technique
Cannabinoids are distinguished from L-trans- cannabinol,
English name (-)-Cannabidiol, which is a very marketable drug
substance. The structural formula of this compound is:
[image]
Currently, cannabidiol is mainly used to protect nerves,
anti-caries, anti-inflammatory and anti-anxiety effects. 2015GW
Biopharmaceuticals announced that it has updated its clinical
trial data on pure cannabinol for refractory childhood epilepsy.
In 2014, the drug has been approved by the US FDA for rare
diseases and fast-track approval for the treatment of infants.
Myoclonic epilepsy, which makes the market prospects for this
drug more widespread. It is cheap, efficient and easy to
operate. It is suitable for the industrial synthetic production
process and will greatly promote the application of cannabidiol.
US20090036523A1 uses olive alcohol as a starting material
and is catalyzed by p-toluenesulfonic acid to obtain the target
product in one step, as follows:
[image]
However, the reaction system is complicated, and there are many
isomers and dimers, and the post-treatment is troublesome. It
requires column chromatography purification and the yield is
low, only 24%, which is not suitable for scale-up production.
WO2006053766A1 utilizes zinc chloride to catalyze the
synthesis of the target product. This document reports a process
for obtaining a product without column purification, but the
purity of the product is only 97.1%, and the yield is only 22%.
We repeated the test on the literature and found that the
largest single impurity in the reaction process is dimer, which
is more than 20%. The impurity needs to be crystallized more
than 3 times to fall within 0.1% (to meet the requirements of
API). When the qualified API is finally obtained, the total
yield is only 13%, and the process cost is high.
US20100298579A1
is prepared by using methyl 2,4-dihydroxy-6-pentanylbenzoate as
a starting material and catalyzed by boron trifluoride diethyl
ether to prepare a coupled methyl ester intermediate (I). The
purity of the reaction is one step. Slightly higher, isomers and
dimers are also significantly less than one-step. However, after
the coupling, the methyl ester intermediate is still only about
75% pure after acid-base treatment, and the melting point of the
compound may be low and cannot be crystallized (intermediate I
has not been reported as melting point, even if the purity of
the column is 98. % of the intermediate I was crystallized and
eventually could not be precipitated as a solid). The methyl
ester intermediate I cannot be recrystallized and purified by a
conventional method, so that the chemical purity and the single
impurity index requirement as key intermediates of the drug
substance cannot be achieved.
[image]
In addition to the above two chemical synthesis methods, there
are some reports that can be obtained by biological extraction
of cannabidiol, but such method steps are cumbersome, and
industrial production limitations are too large.
Summary of
the invention
The object of the present invention is to solve the above
technical problems and to provide a method for synthesizing
cannabidiol.
The object of the invention is achieved by the following
technical solutions:
A method for synthesizing cannabidiol, the reaction formula of
which is as follows
[image]
Including the following steps:
S1, using 2,4-dihydroxy-6-pentylbenzoic acid methyl ester as a
raw material, transesterifying with N,N-dialkylolamine under the
action of potassium hydroxide to obtain intermediate one; the
intermediate Wherein n is 1 to 8, and R is any one of a methyl
group, an ethyl group, a propyl group and a butyl group;
S2, an intermediate prepared by coupling an intermediate 1
obtained in S1 with
(1S,4R)-1-methyl-4-(1-methylvinyl)-2-cyclohexen-1-ol to prepare
an intermediate 2; in the intermediate two, n = 1 to 8, and the
R is any one of a methyl group, an ethyl group, a propyl group
and a butyl group;
S3, the obtained intermediate 2 is hydrolyzed and decarboxylated
under the action of sodium hydroxide to finally obtain
cannabidiol.
Preferably, the S1 comprises the following steps:
S11, using 2,4-dihydroxy-6-pentylbenzoic acid methyl ester as a
raw material and N,N-dialkylolamine, under the action of
potassium hydroxide, reacting by nitrogen gas;
S12, performing the first extraction by adding an acid solution
to adjust the pH of the solution to acidity;
S13, adjusting the pH of the solution to be alkaline by adding a
base, and performing re-extraction;
S14, washed with water, dried, and concentrated under reduced
pressure to give Intermediate 1.
Preferably, the pH of the solution is adjusted to 2-3 in the
S12.
Preferably, the pH of the solution is adjusted to 10 in the S12.
Preferably, the pH is adjusted in the S13 by the addition of a
basic solid.
Preferably, the alkaline solid is added to the S13 as an aqueous
sodium carbonate solid. Of course, the adjustment of the
alkaline alkali solid or the solution can be, the solid is added
to avoid too much water consumption of the alkali, sodium
carbonate, potassium carbonate, cesium carbonate, sodium
hydroxide, potassium hydroxide and other conventional bases can
be.
Preferably, the S2 comprises the following steps:
S21, intermediate one and (1S,
4R)-1-methyl-4-(1-methylvinyl)-2-cyclohexen-1-ol are condensed
under the catalysis of anhydrous zinc chloride;
S22, after the reaction is completed, the acid is added, and the
crude product of the recrystallizable intermediate 2 having a
purity of at least 90% is purified by an acid-base extraction
method;
S23, the crude product of the intermediate 2 is synthesized by
solvent recrystallization to synthesize the high-purity key
intermediate 2 of the bulk drug.
Preferably, the acid in the S22 is any one of hydrochloric acid,
sulfuric acid, acetic acid, oxalic acid and citric acid.
Preferably, the solvent shown in S23 is any one of petroleum
ether, n-pentane, n-hexane and n-heptane.
Preferably, the purity of the cannabidiol produced by the above
method is from 99.88% to 99.98%.
The invention has the beneficial effects that the raw materials
and reagents in the method of the invention are low in price and
commercially available, and high-quality cannabisdiol can be
obtained through a three-step reaction of transesterification,
coupling and decarboxylation. Although the step of the process
is increased to 3 steps, the intermediate of each step can be
purified by recrystallization, and the single impurity can reach
the intermediate index of the raw material medicine, and the
total yield of the qualified raw material medicine can be up to
35~. 40%, the process is obviously improved, and it has a good
industrial application prospect.
Detailed
ways
The method of the present invention will be described below
by way of specific embodiments to make the technical solution of
the present invention easier to understand and grasp, but the
present invention is not limited thereto. The experimental
methods described in the following examples are conventional
methods unless otherwise specified; the reagents and materials
are commercially available unless otherwise specified.
In the embodiment used in the present invention, when pH is
adjusted, sodium carbonate is used in the present application,
but it is not limited thereto, and a conventional base such as
potassium carbonate, cesium carbonate, sodium hydroxide or
potassium hydroxide may be used. Other regulatory auxiliary
agents can also be replaced by other conventional means.
Embodiment
1
Preparation of intermediate one:
First, 119 g of methyl 2,4-dihydroxy-6-pentanebenzoate and
89 g of N,N-dimethylethanolamine were placed in a 250 mL
three-necked flask, and 30.8 g of potassium hydroxide was added
thereto with stirring to protect the nitrogen gas. The
temperature was raised to 130 ° C and the reaction was stirred
for 4 hours. The reaction solution was cooled to below 30 ° C,
adjusted to pH 2-3 with 1N aqueous hydrochloric acid solution,
and extracted with n-heptane (250 mL×2). The aqueous sodium
carbonate solid was adjusted to pH 10, and n-heptane (250 mL×2)
was added. The extract was washed with water (150 mL), dried
over anhydrous sodium sulfate, and evaporated to dryness.
Preparation
of intermediate 2:
100 g of the above transesterified pure product and 800 mL
of dichloromethane were placed in a 2000 mL three-necked flask,
and 50.8 g of zinc chloride, 8 g of water were added under
stirring, and stirred at 25 ° C for 0.5 hour, and 46.4 g of (1S,
4R) was added dropwise.
-1-Methyl-4-(1-methylvinyl)-2-cyclohexen-1-ol, the system is
basically not exothermic, and the mixture is kept warm and
stirred for 24 hours after the completion of the dropwise
addition. The reaction solution was cooled to below 10 ° C,
adjusted to pH 2-3 with 1N aqueous hydrochloric acid solution,
and extracted with n-heptane (500 mL×2). The aqueous sodium
carbonate solid was adjusted to pH 10, and n-heptane (500 mL×2)
was added. The extract was washed with water (300 mL), dried
over anhydrous sodium sulfate and evaporated. The crude product
was added to 4v/m n-heptane at 40 ° C for heating and
dissolution, and the temperature was lowered to -5 to 5 ° C for
crystallization for 16 hours. After suction filtration and
drying, 60.2 g of a white solid was obtained, yield 46%, HPLC
purity 99.98%. .
Synthesis
of cannabidiol:
Add 50g of the above coupled product and 250mL of methanol
to a 1000mL three-necked flask, add 5v/m 3N sodium hydroxide
aqueous solution under nitrogen protection, heat up to 95 °C,
keep the reaction for 8 hours, cool down to 25 °C, add Zhenggeng
The alkane (200 mL*2) was extracted, and the combined organic
phases were washed once with 100 mL of saturated sodium
chloride, concentrated to dryness under reduced pressure, and
then recrystallized from 4 v/m n-heptane, and incubated at -5 to
5 ° C for 16 hours. After filtration and drying, 33.7 g of a
white solid was obtained with a yield of 92% and HPLC purity
99.93%.
Embodiment
2
Preparation of intermediate one:
119 g of methyl 2,4-dihydroxy-6-pentanebenzoate and 234 g of
N,N-dimethylbutanolamine were placed in a 500 mL three-necked
flask, and 33.6 g of potassium hydroxide was added thereto with
stirring to protect the nitrogen gas. The temperature was raised
to 130 ° C and the reaction was stirred for 4 hours. The
reaction solution was cooled to below 30 ° C, adjusted to pH 2-3
with 1N aqueous hydrochloric acid solution, and extracted with
n-heptane (250 mL×2). The aqueous sodium carbonate solid was
adjusted to pH 10, and n-heptane (250 mL×2) was added. The
extract was washed with water (150 mL), dried over anhydrous
sodium sulfate, and evaporated to dryness, and then evaporated
to dryness.
Preparation
of intermediate 2:
100 g of the above transesterified pure product and 800 mL
of dichloromethane were placed in a 2000 mL three-necked flask,
and 46.4 g of zinc chloride, 7.3 g of water were added thereto
with stirring, and stirred at 25 ° C for 0.5 hour, and 42.4 g of
(1S, 4R) was added dropwise.
)-1-methyl-4-(1-methylvinyl)-2-cyclohexen-1-ol, the system is
basically not exothermic, and the mixture is kept warm and
stirred for 24 hours after the dropwise addition. The reaction
solution was cooled to below 10 ° C, adjusted to pH 2-3 with 1N
aqueous sulfuric acid solution, and extracted with n-heptane
(500 mL×2). The aqueous sodium carbonate solid was adjusted to
pH 10, and n-heptane (500 mL×2) was added. The extract was
washed with water (300 mL), dried over anhydrous sodium sulfate
and evaporated. The crude product was added to 4v/m petroleum
ether and dissolved by heating at 40 ° C, and the temperature
was lowered to -5 to 5 ° C for crystallization for 16 hours.
After suction filtration, it was dried to obtain 53.5 g of a
white solid, yield 42%, HPLC purity 99.98%.
Synthesis
of cannabisdiol:
Add 50g of the above coupled product and 250mL of methanol
to a 1000mL three-necked flask, add 5v/m 3N sodium hydroxide
aqueous solution under nitrogen protection, heat up to 95 °C,
keep the reaction for 8 hours, cool down to 25 °C, add Zhenggeng
The alkane (200 mL*2) was extracted, and the combined organic
phases were washed once with 100 mL of saturated sodium
chloride, concentrated to dryness under reduced pressure, and
then recrystallized from 4 v/m n-heptane, and incubated at -5 to
5 ° C for 16 hours. After filtration and drying, 31.3 g of a
white solid was obtained with a yield of 91% and HPLC purity
99.95%.
Embodiment
3:
Preparation of intermediate one:
119 g of methyl 2,4-dihydroxy-6-pentanebenzoate and 145 g of
N,N-dimethylhexanolamine were placed in a 250 mL three-necked
flask, and 30.8 g of potassium hydroxide was added thereto with
stirring to protect the nitrogen gas. The temperature was raised
to 130 ° C and the reaction was stirred for 4 hours. The
reaction solution was cooled to below 30 ° C, adjusted to pH 2-3
with 1N aqueous hydrochloric acid solution, and extracted with
n-heptane (250 mL×2). The aqueous sodium carbonate solid was
adjusted to pH 10, and n-heptane (250 mL×2) was added. The
extract was washed with water (150 mL), dried over anhydrous
sodium sulfate, and evaporated to dryness, and then evaporated
to dryness, and then,
then,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,
Preparation
of intermediate 2:
100 g of the above transesterified pure product and 800 mL
of dichloromethane were placed in a 2000 mL three-necked flask,
and 43.8 g of zinc chloride, 6.9 g of water was added under
stirring, and stirred at 25 ° C for 0.5 hour, and 40 g of (1S,
4R) was added dropwise.
-1-Methyl-4-(1-methylvinyl)-2-cyclohexen-1-ol, the system is
basically not exothermic, and the mixture is kept warm and
stirred for 24 hours after the completion of the dropwise
addition. The reaction solution was cooled to below 10 ° C,
adjusted to pH 2-3 with 1N aqueous sulfuric acid solution, and
extracted with n-heptane (500 mL×2). The aqueous sodium
carbonate solid was adjusted to pH 10, and n-heptane (500 mL×2)
was added. The extract was washed with water (300 mL), dried
over anhydrous sodium sulfate and evaporated. The crude product
was added to 4 v/m of n-hexane at 40 ° C to dissolve by heating,
and the temperature was lowered to -5 to 5 ° C for
crystallization for 16 hours. After suction filtration and
drying, 58.5 g of a white solid was obtained, yield 47%, HPLC
purity 99.91%.
Synthesis
of cannabisdiol:
Add 50g of the above coupled product and 250mL of methanol
to a 1000mL three-necked flask, add 5v/m 3N sodium hydroxide
aqueous solution under nitrogen protection, heat up to 90 °C,
keep the reaction for 4 hours, cool down to 25 °C, add Zhenggeng
The alkane (200 mL*2) was extracted, and the combined organic
phases were washed once with 100 mL of saturated sodium
chloride, concentrated to dryness under reduced pressure, and
then recrystallized from 4 v/m n-heptane, and incubated at -5 to
5 ° C for 16 hours. After filtration and drying, 30.4 g of a
white solid was obtained, yield 94%, HPLC purity 99.88%.
Embodiment 4:
Preparation of intermediate one:
119 g of methyl 2,4-dihydroxy-6-pentanebenzoate and 145.3 g
of N,N-dipropylethanolamine were placed in a 250 mL three-necked
flask, and 30.8 g of potassium hydroxide was added thereto under
stirring to protect with nitrogen. The temperature was raised to
130 ° C and the reaction was stirred for 4 hours. The reaction
solution was cooled to below 30 ° C, adjusted to pH 2-3 with 1N
aqueous hydrochloric acid solution, and extracted with n-heptane
(250 mL×2). The aqueous sodium carbonate solid was adjusted to
pH 10, and n-heptane (250 mL×2) was added. The extract was
washed with water (150 mL), dried over anhydrous sodium sulfate,
and evaporated to dryness, and then evaporated to dryness, and
then,
then,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,
Preparation of intermediate 2:
100 g of the above transesterified pure product and 800 mL
of dichloromethane were placed in a 2000 mL three-necked flask,
and 43.8 g of zinc chloride, 6.9 g of water was added under
stirring, and stirred at 25 ° C for 0.5 hour, and 40 g of (1S,
4R) was added dropwise.
-1-Methyl-4-(1-methylvinyl)-2-cyclohexen-1-ol, the system is
basically not exothermic, and the mixture is kept warm and
stirred for 24 hours after the completion of the dropwise
addition. The reaction solution was cooled to below 10 ° C,
adjusted to pH 2-3 with 1N aqueous sulfuric acid solution, and
extracted with n-heptane (500 mL×2). The aqueous sodium
carbonate solid was adjusted to pH 10, and n-heptane (500 mL×2)
was added. The extract was washed with water (300 mL), dried
over anhydrous sodium sulfate and evaporated. The crude product
was added to 4 v/m of n-heptane at 40 ° C to dissolve by
heating, and the temperature was lowered to -5 to 5 ° C for
crystallization for 16 hours. After suction filtration and
drying, 51 g of a white solid was obtained, yield 41%, HPLC
purity 99.86%.
Synthesis of cannabisdiol:
Add 50g of the above coupled product and 250mL of methanol
to a 1000mL three-necked flask, add 5v/m 3N sodium hydroxide
aqueous solution under nitrogen protection, heat up to 95 °C,
keep the reaction for 8 hours, cool down to 25 °C, add Zhenggeng
The alkane (200 mL*2) was extracted, and the combined organic
phases were washed once with 100 mL of saturated sodium
chloride, concentrated to dryness under reduced pressure, and
then recrystallized from 4 v/m n-heptane, and incubated at -5 to
5 ° C for 16 hours. After filtration and drying, 28.5 g of a
white solid was obtained, yield 88%, HPLC purity 99.97%.
There are many specific embodiments of the invention. All
technical solutions formed by equivalent replacement or
equivalent transformation are within the scope of the claimed
invention.