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Abstract

The paper presents the results of an experimental study regarding the stability of cannabis oil during its long term storage in different conditions. The content of major cannabinoids, namely tetrahydrocannabinol ( Δ–THC), cannabinol (CBN), and cannabidiol (CBD) contained in two batch samples of cannabis oil seizures made by criminal prosecution authorities from Romania was measured during their storage over a period of four years in darkness at 4°C and in laboratory light at 22°C. The results revealed a steadily decay of Δ-THC over the entire storage period from a very high initial content up to a relatively low final content. A slight difference regarding the degree of decay of Δ9–THC between the two storage conditions was recorded, meaning that this is more pronounced when the samples were exposed to light at 22°C. The same trend was recorded for CBD. As expected, the content of CBN increases during storage and the increase is higher when the samples were exposed to light at 22°C.
REV. CHIM. (Bucharest) 63 No. 3 2012 http://www.revistadechimie.ro 293
Long - term Storage and Cannabis Oil Stability
IRENNE GABRIELA TROFIN1*, GABRIEL DABIJA1, DANUT–IONEL VAIREANU2, LAURENTIU FILIPESCU3
1General Inspectorate of Romanian Police, Central Laboratory for Drug Analysis and Profiling, Bucharest, Romania.
2University Politehnica of Bucharest, Faculty of Applied Chemistry and Materials Science, Department of Applied Physical
Chemistry and Electrochemistry, 313 Splaiul Independentei, 060042, Bucharest, Romania
3University Politehnica of Bucharest, Faculty of Applied Chemistry and Materials Science, Department Technology of Inorganic
Substances and Environmental Protection, 313 Splaiul Independentei, 060042, Bucharest, Romania
The paper presents the results of an experimental study regarding the stability of cannabis oil during its long-
term storage in different conditions. The content of major cannabinoids, namely tetrahydrocannabinol (
Δ
9
THC), cannabinol (CBN), and cannabidiol (CBD) contained in two batch samples of cannabis oil seizures
made by criminal prosecution authorities from Romania was measured during their storage over a period of
four years in darkness at 4°C and in laboratory light at 22°C. The results revealed a steadily decay of
Δ
9–THC
over the entire storage period from a very high initial content up to a relatively low final content. A slight
difference regarding the degree of decay of
Δ
9–THC between the two storage conditions was recorded,
meaning that this is more pronounced when the samples were exposed to light at 22°C. The same trend was
recorded for CBD. As expected, the content of CBN increases during storage and the increase is higher when
the samples were exposed to light at 22°C.
Keywords: cannabis, oil, decay, cannabinoid
Cannabis oil, often called hashish oil, is a liquid cannabis
product with a high content of tetrahydrocannabinol (Δ9
THC) obtained by extraction from either herbal cannabis
or from cannabis resin. Although such a product requires
more complex preparation methods than other cannabis
products, it is often preferred by drug dealers because they
may traffic more psychoactive material in a smaller
quantity of cannabis product [1].
As with other cannabis products, cannabis oil has
different stability in various environmental conditions [2-
4]. The psychoactive component, namely Δ9THC, has a
relatively high instability when the cannabis products are
exposed to air, light or acidic environments [5-7]. Its
instability to heat has been also demonstrated [8]. It is
widely accepted that the main pathway of cannabis
products deactivation is the conversion of Δ9–THC to
cannabinol (CBN). After five years of storage in an ethanol/
propylene glycol solution of a Δ9–THC sample, it was
proposed a correlation between the oxidative derivatives
of this cannabinoid formed during its degradation to the
less psychoactive component CBN [9].
* email: irennetrofin@yahoo.com
REV. CHIM. (Bucharest) 63 No.3 2012http://www.revistadechimie.ro294
Other major cannabinoid present in cannabis products, namely cannabidiol may also suffer changes depending on
storage conditions. In this respect, under acidic conditions it may transform into Δ9–THC by acid-catalyzed cyclization
and, in the presence of oxygen, is oxidized to monomeric and dimeric hydroxyquinones [10].
Although the cannabinoids stability was intensively
studied previously, most of the researches were done on
pure solutions, which can have a different behaviour
towards actual cannabis products. Therefore, the objective
of this paper is to explore experimentally the influence of
storage conditions such as temperature and light on the
stability of the major cannabinoids in the cannabis oil.
Experimental part
Chemicals and reagents
All chemicals and reagents used for samples preparation
and analysis were of analytical grade from Merck
(Darmstadt, Germany). The etalons of Δ9–tetrahydro-
cannabinol (Δ9–THC), cannabidiol (CBD), and cannabinol
(CBN) were purchased from Switzerland. The ultrapure
water used in HPLC analyses was prepared in-house using
a Millipore system, model Milli-Q Integra 3.
Cannabis oil samples
Cannabis oil from two different seizures (marked with
symbols U1 and U2) made by criminal prosecution
authorities from Romania and provided by Central
Laboratory for Drug Analysis and Profiling were subject to
experimental investigation. The samples of cannabis oil
were supplied in small bottles and have a black-brown
colour and a high consistency comparable to that of a paste.
The samples were stored in the darkness at 4°C and in the
laboratory light at 22°C for four years. At regular intervals,
namely at every three months, samples were taken for
analysis in order to determine the content of their major
cannabinoids (Δ9THC, CBD, and CBN).
Methods
The procedure that led to the sample preparation for
analysis consisted of extracting 0.05 g of cannabis oil in 20
mL of a methanol-chloroform (9:1, v/v) mixture. Thus, the
samples were shaken for 30 min and then placed in an
ultrasonic bath at ambient temperature for 15 min in order
to increase the cannabinoids extraction rates. The extracts
were filtered and some aliquots (0.6 mL) of the filtrates
were transferred to a 4 mL vials and then evaporated to
dryness by oven evaporation, only up to 80°C for prevention
of any decomposition reactions. Then, the vials were put
into a heating unit at 220°C for 12 min when the traces of
tetrahydrocannabinolic acid (THCA) are decarboxylated.
Decarboxylation is particularly required for the
determination of the entire content of Δ9THC of the
sample. Before analyses, the residues were extracted in
1.5 mL extraction solvent (methanol-chloroform 9:1, v/v).
After this, the samples were subject to analyses of the
major cannabinoids content (Δ9THC, CBD, and CBN) [1].
Analytical protocol
Extracts obtained by procedure described above have
been subject to analytical investigations through
instrumental methods (GCFID – Gas Chromatography
Flame Ionization Detector and HPLC – High Performance
Liquid Chromatography) in order to find out the content in
major cannabinoids (Δ9THC, CBD, and CBN).
GCFID analyses were carried out on a 7890A gas
chromatograph with a flame ionization detector.
Separation was achieved on a fused silica capillary column
(HP-5MS, 30 m×0.32 mm i.d., 0.25μm film thickness, J&W
Scientific, Folsom, CA, USA). Temperature program: 150°C
hold for 1 min, 10°C/min to 280°C, hold for 5 min. The
injection port and interface temperature were 250°C and
300°C, respectively. Split injection mode was used (20:1)
and hydrogen, with a flow rate of 30 mL per min, was used
as carrier gas [11].
HPLC analyses were carried out on an Agilent 1100
Series HPLC chromatograph equipped with a quaternary
pump, autosampler, column oven and diode-array detector
(DAD) UV Lamp ON (223 nm). Chromatography was
achieved on a 250 mm × 4.6 mm i.d., 5 μm Hypersil ODS
column. The HPLC operates with constant flow at 1 mL
mobile phase (acetonitrile 37.5% and ultrapure water) per
minute.
Results and discussions
GCFID and HPLC chromatograms revealed a very high
content of ΔΔ
ΔΔ
Δ9THC in the cannabis oil samples and
subsequently, a very high potency of these type of cannabis
product compared to other cannabis products such as
herbal cannabis and cannabis resin. The difference
between the two samples regarding their potency is
significant and comes probably from the use of different
procedures for their obtaining. The difference could be due
to the type of solvent used for extraction, number of
extractions, conditions of extractions etc. These results
indicate, on the one hand, that the samples were prepared
in different clandestine laboratories, improvised by different
drug traffickers and, on the other hand, that the samples
went through different routes of trafficking originating from
different geographical areas. Following a detailed analysis
of such data, the prosecution authorities could identify
trafficking routes, and finally the places where this type of
drugs were made.
REV. CHIM. (Bucharest) 63 No. 3 2012 http://www.revistadechimie.ro 295
The experimental results concerning the stability of the
major cannabinoids indicate a small but constant
difference between cannabinoids content of the cannabis
oil depending on storage conditions. Figures 1 and 2 show
the variation of the major cannabinoids content in the two
samples of cannabis oil as a function of time and storage
conditions. As one can see, in both samples the Δ9THC
content decreases during storage and is always higher in
the samples stored in the darkness at 4°C than in the
samples stored in the laboratory light at 22°C. The same
trend can be also observed for CBD content. As it was
expected, the CBN content increases during storage period
and is always higher in the samples stored in laboratory
light at 22°C than in the samples stored in darkness at 4°C.
The results obtained for the two samples derived from the
two different seizures regarding the evolution of the major
cannabinoids content during storage in different conditions
are presented in table 2. The results revealed a steadily
decay of Δ9THC over the entire storage period. Moreover,
the decay of Δ9THC in the samples exposed to light at
22°C is higher than in the samples stored in the darkness at
4°C. In this respect, when the samples from seizure U1
were stored in the darkness at 4°C, 21.6% of Δ9THC (fig. 1
a) was lost in the first year with an average loss of 4.4%
every tree months, 21.83% in the second year with an
average loss of 5.46%, 21.64% in the third year with an
average loss of 5.41%, and 18.69% in the fourth year with
an average loss of 4.67%. When the samples from the same
seizure were stored in the laboratory light at 22°C, 23.16%
of Δ9THC was lost in the first year with an average loss of
5.79% every tree months, 23.25% in the second year with
an average loss of 5.81%, 22.99% in the third year with an
average loss of 5.75%, and 20.45% in the fourth year with
an average loss of 5.11%. Finally, after four years of storage,
the samples stored in the darkness at 4°C lost 83.75% of
Δ9THC and the samples stored in the laboratory light at
22°C lost 89.85% of Δ9THC ( 6.1% higher).
The variation of CBN (fig. 1 b) of the same samples over
the storage period indicates that when these were stored
in the darkness at 4°C, 59.03% of CBN was formed in the
first year with an average gain of 14.76% every three
months, 16.49% in the second year with an average gain
of 4.12%, 4.76% in the third year with an average gain of
1.19%, and 3.01% in the fourth year with an average gain of
0.75%. When the samples from the same seizure were
stored in the laboratory light at 22°C, 65.29% of CBN was
formed in the first year with an average gain of 16.32%
every three months, 14.18% in the second year with an
average gain of 3.54%, 3.72% in the third year with an
Table 1
THE INITIAL CONTENT OF MAJOR CANNABINOIDS
IN CANNABIS OIL
Table 2
EVOLUTION OF THE MAJOR CANNABINOIDS CONTENT IN
CANNABIS OIL DURING STORAGE IN DIFFERENT CONDITIONS
Fig. 1 Variation of cannabinoids content in
cannabis oil: (a) Δ9–THC decay;
(b) CBN formation; (c) CBD decay
REV. CHIM. (Bucharest) 63 No.3 2012http://www.revistadechimie.ro296
Fig. 2 Decay degree of Δ9–THC in
cannabis oil derived from:
(a) seizure U1; (b) seizure U2
average gain of 0.93%, and 3.95% in the fourth year with an
average gain of 0.99%. Finally, after four years, the samples
stored in the darkness at 4°C gained 83.29% of CBN and
the samples stored in the laboratory light at 22°C gained
87.15% of CBN (3.86% higher). The same trend was also
recorded for the second cannabis oil sample.
Comparing the decay degree of Δ9–THC in the first year
of storage period of cannabis oil with the formation degree
of CBN in the same year, it can be seen that the latter yield
is with about 35% higher than the first yield. In this respect,
the changes regarding the content of CBN during the
storage period can not be entirely correlated with the
chemical and/or biochemical decay processes of Δ9–THC
to CBN. Some other unknown variables beyond the control,
such as the origin place, degradation already started during
the trafficking transports play also an important role. Also,
other degrading routes of other cannabinolic compounds
must be considered as contributors to the overall increase
of CBN content upon long-term storage.
The variation of CBD (fig. 1 c) in the same samples over
the entire storage period indicates that in the case of
samples from seizure U1 stored in the darkness at 4°C,
11.03% of CBD was lost in the first year with an average
loss of 2.76% every tree months, 12.15% in the second
year with an average loss of 3.04%, 7.47% in the third year
with an average loss of 1.87%, and 9.54% in the fourth year
with an average loss of 2.39%. When the samples from the
same seizure were stored in the laboratory light at 22°C,
13.45% of CBD was lost in the first year with an average
loss of 3.35% every three months, 12.05% in the second
year with an average loss of 3.01%, 6.81% in the third year
with an average loss of 1.7%, and 8.5% in the fourth year
with an average loss of 2.12%. Finally, after four years of
storage, the samples stored in the darkness at 4°C lost
40.18% of CBD and the samples stored in the laboratory
light at 22°C lost 44.8% of CBD (with less 0.62%). The same
trend was recorded for all cannabis oil samples.
Analyzing these results it can be seen that the decay
degree of CBD in the first year of storage period of cannabis
oil is about 10% (approximately quarter the difference of
35% between the decay degree of Δ9–THC and respectively,
the formation degree of CBN) in the case of the samples
stored in the darkness at 4°C and about 19%
(approximately half the difference of 40% between the
decay degree of Δ9–THC and respectively the formation
degree of CBN) was observed in the case of the samples
stored in the laboratory light at 22°C. These results suggest
a different degrading rout of CBD, probably dependent on
the storage conditions. When the samples were stored in
the darkness one of the degradative route could be the
biochemical cyclization of CBD to Δ9–THC, followed by
the oxidative decay of Δ9–THC to CBN. In addition, when
the samples were exposed to light, CBD might achieve
photo-reactive properties and transforms into Δ9–THC [10].
A pseudo zero-order kinetic was used (fig. 3) in order to
calculate the kinetic parameters of the Δ9–THC decay such
as the rate constant (k), the half-time (t1/2), and the decay
rate (v). The linear regression parameters and the
correlation coefficients are presented in the table 3. As
can be seen from the table 4, both rate constant and decay
rate are higher in the samples stored in the laboratory light
at 22°C than those stored in the darkness at 4°C. The values
of the half-time corresponding to the samples stored in the
laboratory light at 22°C are smaller than those stored in the
darkness at 4°C. These results suggest a higher rate of Δ9
THC decay in the cannabis oil stored in normal conditions
(natural light and ambiental temperature) than in the case
of special storage conditions (darkness and low
temperature).
Fig. 3 Pseudo zero-order kinetic of Δ9–THC decay in the cannabis oil; the solid line represents the linear regression of data
corresponding to 4oC and darkness storage conditions and, the dashed line represents the linear regression of data corresponding
to 22oC and laboratory light storage conditions
REV. CHIM. (Bucharest) 63 No. 3 2012 http://www.revistadechimie.ro 297
Table 4
KINETIC PARAMETERS OF Δ9-THC DECAY
CALCULATED FROM A PSEUDO-ZERO
ORDER KINETIC
Table 3
LINEAR REGRESSION PARAMETERS
AND CORRELATION COEFFICIENTS
Conclusions
Chemical characterization of the cannabis oil samples
derived from two different seizures revealed a very high
content of Δ9–THC compared with other cannabis products
such as the herbal cannabis and cannabis resin. The
difference between the potency of the two samples
suggests a different preparation method or a different route
of trafficking, which could finally indicate the places where
the samples were prepared.
The experimental results regarding the stability of the
major cannabinoids species revealed differences that
should be taken into consideration between cannabinoids
content of the cannabis oil are related to the storage
conditions. Thus, the results revealed a steady decay of
Δ9–THC over the entire storage period. Moreover, the decay
of Δ9–THC in the samples exposed to light at 22°C is more
pronounced than in the samples stored in the darkness at
4°C. The content of CBN increases during storage, and
increase is more pronounced for the samples exposed to
light at 22°C than those stored in the darkness at 4°C. These
results are in part consistent with those obtained for Δ9
THC. The CBD content decreases during storage especially
for samples exposed to light at 22°C. This evolution could
be explained by considering the biochemical cyclization
of CBD to Δ9–THC, followed by the decay of Δ9–THC to CBN
as a degrading route for the samples stored in the darkness
at 4°C and, both biochemical and photochemical
cyclization of CBD to Δ9–THC followed by decay of Δ9–THC
to CBN as a degrading routes for the samples exposed to
light at 22°C.
The decay of Δ9-THC takes place up on a pseudo zero-
order kinetic and the calculated values of the kinetic
parameters suggest a higher rate of Δ9–THC decay in
normal storage conditions such as light and ambiental
temperature than in special ones such as darkness and
low temperature.
References
1. *** United Nations Office on Drugs and Crime (UNODC), Manual
for use by national drug analyses laboratories, 2009.
2. MARTONE G., DELLA CASA E., Forensic. Sci. Int., 47, 1990, p. 147.
3. LINDHOLST C., AUSTRALIAN J. Forensic. Sci., 42, no 3, 2010, p. 181.
4. ROSS S. A., ELSOHLY M. A., Bull. Narc., 49, no.50, 1997, p. 139.
5. MECHOULAM R., HANUŠ L., Chem. Phys. Lipids., 108, 2000, p. 1.
6. GOLDING F.S., DÍAZ-FLORES E.J., DÍAZ R.C., Ann. Clin. Lab. Sci.,
28, no.3, 1998, p. 160.
7. DUSSY F.E., HAMBERG C., LUGINBÜHL M., SCHWERZMANN T.,
BRIELLMANN T.A., Forensic Sci. Int., 149, 2005, p. 3.
8. TROFIN I.G., VLAD C.C., DABIJA G., FILIPESCU L., Rev.
Chim.(Bucharest), 62, no. 6, 2011, p. 639
9. CARBONE M., CASTELLUCCIO F., DANIELE A., SUTTON A ., LIGRESTI
A., DI MERZO V., GAVAGNIN M., Tetrahedron, 66, no. 49, 2010, p. 9497.
10. MECHOULAM R., HANUŠ L., Chem. Phys. Lipids., 121, 2002, p. 35.
11. VLAD C.C., TROFIN I.G., DABIJA G., FILIPESCU L., Rev. Chim.
(Bucharest), 62, no. 9, p. 916
Manuscript received: 16.08.2011
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Cannabis is one of the oldest cultivated plant, which has been used by humankind for thousands of years due to its biological properties and a wide range of applications. In total, hemp plants contain over 500 different substances while the characteristic components are the cannabinoids. The most important cannabinoids are (-)-Δ⁹-trans-tetrahydrocannabinol (Δ⁹-THC), cannabidiol (CBD), and cannabinol (CBN – the latter being an oxidation product resulting from Δ⁹-THC). In the course of recent years, a paradigm shift has taken place with regard to the use of products and ingredients derived from hemp, especially CBD. Thus, an ever-increasing number of products containing CBD are on the market; this ranges from classic CBD oil to CBD chewing gum and even CBD shampoo. Despite an increasing presence of these products in the market, the regulation of cannabinoids in these products is very inconsistent in different countries, except for Δ⁹-THC whose limit is 0.2% for many products and many countries. The enormous abundance of CBD-containing products calls for the development of new analytical techniques that allow a reliable and quick determination of the main cannabinoids usually found in hemp. This seems all the more necessary since previous examinations of CBD oils often revealed a difference between the declared amount and the actual content of the ingredients. Many methods usually applied to determine cannabinoids are rather time-consuming and associated with high costs. In this study, we developed and validated a sensitive, simple, reliable as well as fast method for the determination of CBN, CBD and Δ⁹-THC in commercially available CBD oils using high-performance thin-layer chromatography (HPTLC) combined with electrospray ionization mass spectrometry (ESI-MS). Thus, for this method, a recovery rate of ≥90% was determined. This procedure enables both qualitative and quantitative analyses of CBN, CBD and Δ⁹-THC in CBD oils of different matrices such as hempseed oil, olive oil or sunflower oil. Thus, this method is a helpful and fast tool to investigate a broad variety of commercially available CBD oils.
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Background Cannabis has been used worldwide for centuries for industrial, recreational and medicinal use, however, to date no successful attempts at editing genes involved in cannabinoid biosynthesis have been reported. This study proposes and develops an in silico best practices approach for the design and implementation of genome editing technologies in cannabis to target all genes involved in cannabinoid biosynthesis. Results A large dataset of reference genomes was accessed and mined to determine copy number variation and associated SNP variants for optimum target edit sites for genotype independent editing. Copy number variance and highly polymorphic gene sequences exist in the genome making genome editing using CRISPR, Zinc Fingers and TALENs technically difficult. Evaluation of allele or additional gene copies was determined through nucleotide and amino acid alignments with comparative sequence analysis performed. From determined gene copy number and presence of SNPs, multiple online CRISPR design tools were used to design sgRNA targeting every gene, accompanying allele and homologs throughout all involved pathways to create knockouts for further investigation. Universal sgRNA were designed for highly homologous sequences using MultiTargeter and visualised using Sequencher, creating unique sgRNA avoiding SNP and shared nucleotide locations targeting optimal edit sites. Conclusions Using this framework, the approach has wider applications to all plant species regardless of ploidy number or highly homologous gene sequences. Significance statement Using this framework, a best-practice approach to genome editing is possible in all plant species, including cannabis, delivering a comprehensive in silico evaluation of the cannabinoid pathway diversity from a large set of whole genome sequences. Identification of SNP variants across all genes could improve genome editing potentially leading to novel applications across multiple disciplines, including agriculture and medicine.
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The aim of this work was to validate a high-performance liquid chromatography (HPLC) method for determination of the stability of cannabidiol, ∆9-tetrahydrocannabinol, and cannabinol. Furthermore, degradation kinetics were also investigated. Five stress conditions—acid degradation, alkaline degradation, oxidation, thermal degradation, and photodegradation—were evaluated. The results showed that the HPLC method had a linear response (R2 ≥ 0.9999) in the test range of 1–200 μg/mL. The method was specific, precise, and accurate. The limits of both detection and quantitation are also reported. According to the stress test, the three cannabinoids (cannabidiol, ∆9-tetrahydrocannabinol, cannabinol) were stable during exposure to a range of thermal conditions for 24 h. They were unstable when being subjected to acid conditions; cannabidiol under alkaline conditions was extremely unstable. Degradation kinetic analysis demonstrated that the compounds remained at a level of approximately 8% after 5 h, and approached a first-order reaction (R2 = 0.9930) with a rate constant of − 0.5057 h− 1. In summary, the obtained data can be used as a guide for the formulation development of cannabis products in order to maintain their active compounds as well as their activities.
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Over the last few years considerable attention has focused on cannabidiol (CBD), a major non-psychotropic constituent of Cannabis. In Part I of this review we present a condensed survey of the chemistry of CBD; in Part II, to be published later, we shall discuss the anti-convulsive, anti-anxiety, anti-psychotic, anti-nausea and anti-rheumatoid arthritic properties of CBD. CBD does not bind to the known cannabinoid receptors and its mechanism of action is yet unknown. In Part II we shall also present evidence that it is conceivable that, in part at least, its effects are due to its recently discovered inhibition of anandamide uptake and hydrolysis and to its anti-oxidative effect.
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The concentration of Δ9-tetrahydrocannabinol (THC) and cannabinol (CBN) in cannabis plant material (marijuana) of different varieties stored at room temperature (20-22°Celsius (C)) over a four-year period was determined. The percentage loss of THC was proportional to the storage time. On average, the concentration of THC in the plant material decreased by 16.6% ±7.4 of its original value after one year and 26.8% ±7.3, 34.5% ±7.6 and 41.4% ±6.5 after two, three and four years, respectively. A relationship between the concentration ratio of CBN to THC and the storage time was developed and could serve as a guide in determining the approximate age of a given marijuana sample stored at room temperature.
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The aim of the present paper was to investigate the stability of cannabinoids in herbal cannabis upon long-term storage. The content of tetrahydrocannabinol (Δ 9 -THC), cannabinol (CBN), and cannabidinol (CBD) in herbal cannabis from ten different regions of the world were measured for up to four years of storage in darkness at 4°C and in natural light of laboratory at 22°C. The degradation of Δ 9 -THC was faster in the first year than in subsequent years, and more pronounced for the samples exposed to light at 22°C than those stored in darkness at 4°C. The content of CBN increases during the storage and the increase is more pronounced for the samples exposed to light at 22°C than those stored in the darkness at 4°C. These results are consistent with those obtained for Δ 9 -THC. Also, a new criterion for the chemical potency ranking in different herbal cannabis grades was approached on the basis of the Δ 9 -THC degradation kinetics.
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The chemical analysis of a sample of Δ9-THC, which had been stored in an ethanol/propylene glycol solution for 5 years, resulted in the isolation of several hydroxylated Δ9-THC derivatives, the main of which were trans-cannabitriol monoethyl ether (4) and trans-propanediol ethers 7 and 8. cis-Cannabitriol monoethyl ether (5) and the oxidised derivatives 3 and 6 were detected in lesser amounts. The structure elucidation of the unprecedented cannabinoids 3, 5, 7 and 8 was achieved mainly by NMR techniques. Full NMR assignment of compounds 4 and 6 were also made. The detection of cannabitriol (6) and the corresponding solvent-adduct analogues (compounds 4–8) was in agreement with the decomposition mechanisms previously proposed for Δ9-THC. The isolation of the endoperoxide 3 represents indirect evidence of the existence of unstable precursors that were suspected to be intermediates in the non-enzymatic oxidation pathway of Δ9-THC. Both isomers of cannabitriol monoethyl ether exhibited weak affinity at either CB1 (Ki=2.25, 6.30 μM) or CB2 cannabinoid receptors (Ki=1.97, 3.13 μM), the trans isomer always being more potent than the cis isomer.
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The aim of the present study was to investigate the stability of cannabinoids in cannabis resin slabs and cannabis extracts upon long-term storage. The levels of tetrahydrocannabinol (THC), cannabinol (CBN), cannabidiol (CBD) and cannabigerol (CBG) on both neutral and acidic form were measured at room temperature, 4°C and −20°C for up to 4 years. Acidic THC degrades exponentially via decarboxylation with concentration halve-lives of approximately 330 and 462 days in daylight and darkness, respectively. The degradation of neutral THC seems to occur somewhat slower. When cannabinoids were stored in extracted form at room temperature the degradation rate of acidic THC increased significantly relative to resin material with concentration halve-lives of 35 and 91 days in daylight and darkness, respectively. Once cannabis material is extracted into organic solvents, care should be taken to avoid the influence of sunlight.
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Hashish samples from Morocco, Lebanon, Iran, India, Pakistan, Afghanistan have been stored either in darkness or in daylight at 18–22°C and analysed every 20–30 days for a period of 22 months. Each of the samples was analysed by gas-chromatography with α-cholestane as the internal standard. As a result of this study we have observed distinctive ageing processes due to the physical and chemical peculiarity of samples of different geographic origin. The information obtained from this work may have a forensic application in the illicit traffic of hashish.
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The chemical research on the plant cannabinoids and their derivatives over two centuries is concisely reviewed. The tortuous path leading to the discovery of the endogenous cannabinoids is described. Future directions, which will probably be followed are delineated.
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A simple procedure based on a common silica gel column chromatography for the isolation of Delta9-tetrahydrocannabinolic acid A (Delta9-THCA-A) from hemp in a multi-milligram scale is presented. Further, the decarboxylation reaction of Delta9-THCA-A to the toxicologically active Delta9-tetrahydrocannabinol (Delta9-THC) at different analytical and under-smoking conditions is investigated. Maximal conversion in an optimised analytical equipment yields about 70% Delta9-THC. In the simulation of the smoking process, only about 30 % of the spiked substance could be recovered as Delta9-THC.
  • Martone G Della
MARTONE G., DELLA CASA E., Forensic. Sci. Int., 47, 1990, p. 147.
  • Lindholst C Australian
LINDHOLST C., AUSTRALIAN J. Forensic. Sci., 42, no 3, 2010, p. 181.