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Terpenes in Cannabis sativa – From plant genome to humans



Cannabis sativa (cannabis) produces a resin that is valued for its psychoactive and medicinal properties. Despite being the foundation of a multi-billion dollar global industry, scientific knowledge and research on cannabis is lagging behind compared to other high-value crops. This is largely due to legal restrictions that have prevented many researchers from studying cannabis, its products, and their effects in humans. Cannabis resin contains hundreds of different terpene and cannabinoid metabolites. Many of these metabolites have not been conclusively identified. Our understanding of the genomic and biosynthetic systems of these metabolites in cannabis, and the factors that affect their variability, is rudimentary. As a consequence, there is concern about lack of consistency with regard to the terpene and cannabinoid composition of different cannabis ‘strains’. Likewise, claims of some of the medicinal properties attributed to cannabis metabolites would benefit from thorough scientific validation.
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Title: Terpenes in Cannabis sativa – From plant genome to
Authors: Judith K. Booth, J¨org Bohlmann
PII: S0168-9452(19)30119-0
Reference: PSL 10109
To appear in: Plant Science
Received date: 26 January 2019
Revised date: 27 March 2019
Accepted date: 27 March 2019
Please cite this article as: Booth JK, Bohlmann J, Terpenes in
Cannabis sativa – From plant genome to humans, Plant Science (2019),
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Terpenes in Cannabis sativa From plant genome to humans
Judith K. Booth1 and Jörg Bohlmann1
1Michael Smith Laboratories, University of British Columbia, 2185 East Mall, Vancouver, B.C., Canada,
V6T 1Z4
Author for Correspondence: Dr. Jörg Bohlmann
Michael Smith Laboratories, University of British Columbia, 2185 East Mall, Vancouver, B.C., Canada
Highlights: In this Review Article we highlight recent progress in research on terpene metabolites in
cannabis. This includes highlighting issues related to claims of medicinal properties of cannabis terpenes.
We also highlight issues related to so-called ‘strains’ and the need to establish properly genotyped
cannabis varieties for research and use in the industry, as well as other areas that require future
Table of contents
3 Introduction
4 Chemistry, biosynthesis, and genomics of terpene diversity and variation in cannabis
6 Biosynthesis of cannabinoids
7 Effects attributed to terpenes in cannabis
7 Claims of anticancer effects of cannabis and cannabis terpenes may do more harm than good
8 Perspectives and future directions
Cannabis sativa (cannabis) produces a resin that is valued for its psychoactive and medicinal properties.
Despite being the foundation of a multi-billion dollar global industry, scientific knowledge and research
on cannabis is lagging behind compared to other high-value crops. This is largely due to legal restrictions
that have prevented many researchers from studying cannabis, its products, and their effects in humans.
Cannabis resin contains hundreds of different terpene and cannabinoid metabolites. Many of these
metabolites have not been conclusively identified. Our understanding of the genomic and biosynthetic
systems of these metabolites in cannabis, and the factors that affect their variability, is rudimentary. As
a consequence, there is concern about lack of consistency with regard to the terpene and cannabinoid
composition of different cannabis strains. Likewise, claims of some of the medicinal properties
attributed to cannabis metabolites would benefit from thorough scientific validation.
1. Introduction
Cannabis sativa (cannabis) is thought to have originated from central Asia, and has been domesticated
for over 5,000 years [1]. Cannabis varieties that are low in psychoactive cannabinoids are used for the
production of fiber and oilseed. However, the most valuable cannabis product today is the terpene- and
cannabinoid-rich resin with its various psychoactive and medicinal properties. The resin is produced and
accumulates in glandular trichomes that densely cover the surfaces of female (pistillate) inflorescences
and, to a lesser degree, the foliage of male and female plants (Figure 1). In total, more than 150
different terpenes and approximately 100 different cannabinoids [2] (Figure 2) have been identified in
the resin of different cannabis types (Table 1). The predominant cannabinoids in cannabis grown for
medicinal or recreational use are Δ9-tetrahydrocannabinolic acid (THCA) and cannabidiolic acid (CBDA).
While cannabinoids are the primary psychoactive and medicinal components of cannabis resin, volatile
terpenes (monoterpenes and sesquiterpenes) contribute many of the different fragrance attributes that
influence consumer preferences.
Different cannabis types and their derived consumer products are commonly referred to with strain
names. These names often relate to fragrance attributes conferred, at least in part, by terpenes [3].
Different strains may be distinguished by morphological features or differences in the chemical
composition of the resin. However, due to a history of largely illicit cannabis production, cannabis
strains are often poorly defined genetically. Strains may lack reproducibility with regard to profiles of
terpenes and cannabinoids [4, 5]. The species encompasses large genetic diversity, with most strains
having high levels of heterozygosity and genetic admixture [5, 6]. Cannabis is wind-pollinated, which also
contributes to variability of cannabis metabolites. As a result, many cannabis strains lack the level of
standardization that producers and consumers are accustomed to with other crop plants, such as
genetically and phenotypically well-defined grapevine varieties. In the absence of proper genetic or
genomic characterization, some attempts have been made at chemotaxonomic classification of cannabis
strains based on terpenes, and cannabis plants have also been described as belonging to different
chemotypes (Table 1). However, the complexity of terpene biosynthetic systems, and the many
different sources of terpene variation, renders these efforts often futile; in general, concepts of
chemotaxonomy have been outdated by genome sciences, and chemotypes cannot reliably substitute
for properly genotyped plants.
With the lifting of some of the legal restrictions on cannabis research in Canada, and in some other
jurisdictions, there is now an opportunity to build stronger scientific knowledge of the genomic,
molecular and biochemical properties that define terpene and cannabinoid profiles in different cannabis
strains. This in turn can support the development of a larger number of well-defined cannabis varieties.
Another aspect that requires new research are the various effects that are attributed to cannabis
terpenes in humans. While some of the effects of the cannabinoids have been scientifically explained,
there is a great deal of uncertainty about the effects of cannabis terpenes in humans beyond fragrance
2. Chemistry, biosynthesis and genomics of terpene diversity and variation in cannabis
Terpene composition is a phenotypic trait that shows much variation across different cannabis strains
(Table 1). The majority of terpenes found in cannabis are hydrocarbons, which are the direct products of
terpene synthase (TPS) enzymes [7, 8], as opposed to more complex terpenes that require modification
by other enzymes such as cytochrome P450s. Therefore, the chemical diversity of cannabis terpenes
reflects the diversity of TPS enzymes encoded in the cannabis (Cs)TPS gene family.
The monoterpene myrcene as well as the sesquiterpenes β-caryophyllene and α-humulene appear to be
present in most cannabis strains. Other common compounds include the monoterpenes α-pinene,
limonene, and linalool as well as the sesquiterpenes bisabolol and (E)-β-farnesene. It is important to
note that some terpenes, in particular sesquiterpenes, remain difficult to identify due to the lack of
authentic standards for many of these compounds. As a result, reports of terpene profiles in cannabis
may include unknown compounds, rely on tentative identification, or present incomplete profiles of
selected compounds. Stereochemistry is also not consistently described, or is often ignored, in reports
on cannabis terpenes. These issues makes it difficult to fully assess the diversity of terpenes in cannabis
using the available data and make it problematic to compare the results of different studies.
The terpenes found in the cannabis resin, as well as the isoprenoid moiety of the cannabinoid structure,
are produced through the isoprenoid biosynthetic system, which originates in the mevalonic acid (MEV)
pathway in the cytosol and the methylerythritol phosphate (MEP) pathway in plastids. Monoterpenes
and cannabinoids have a common ten-carbon isoprenoid precursor, geranyl diphosphate (GPP, C10),
while sesquiterpenes are produced from the fifteen-carbon isoprenoid farnesyl diphosphate (FPP, C15).
Using GPP or FPP as substrates, monoterpene synthases (mono-TPS) and sesquiterpene synthases
(sesqui-TPS) produce the different structures of mono- and sesquiterpenes found in the cannabis resin
(Figure 2). A recent analysis of the Purple Kush cannabis genome and transcriptome sequences
identified more than 30 different CsTPS genes [8]. Only nine CsTPS have been functionally characterized
and published to date [8, 9]. As with many other plant TPS [7], eight of the nine characterized CsTPS are
multi-product enzymes that generate several different terpene structures from either GPP or FPP [8].
The multi-product nature of CsTPS can explain why some terpenes, such as α-humulene and β-
caryophyllene, typically co-occur in different cannabis samples. The CsTPS responsible for many of the
different terpenes found in cannabis are still unknown.
Variation of the composition of the CsTPS gene family and variation in CsTPS gene expression is likely to
explain observed variations of terpene profiles across the species. However, the level of variation of the
size, composition and expression of the CsTPS gene family, and factors that influence CsTPS gene
expression, are for the most part unknown. For example, variation of terpene biosynthesis at the
genome, transcriptome, proteome and biochemical levels have been shown in other plants to account
for phenotypic intra-specific variation of terpene profiles [eg. 10, 11]. Terpene profiles may also
substantially change as a result of differential CsTPS gene expression over the course of plant
development or in response to environmental factors. In addition, developmental or tissue specific
expression of CsTPS may affect variation of terpene profiles in cannabis products. None of these factors
of terpene variation, which may contribute to poor reproducibility of terpene composition, have been
systematically studied in cannabis.
The oxygen functionality of simple terpene alcohols found in cannabis such as linalool or bisabolol may
result from the enzymatic activity of CsTPS as has also been shown for TPS in other plants species [8].
Other terpene derivatives detected in cannabis may arise non-enzymatically due to oxidation or due to
thermal- or UV-induced rearrangements during processing or storage, such as caryophyllene oxide, β-
elemene, or derivatives of myrcene [8, 12]. These non-enzymatic modifications may add a level of
variation that is independent of the plant genome and biochemistry. When terpene analysis is
performed with dried plant material, variable quantitative losses of terpenes, especially the more
volatile monoterpenes [13], may be another cause of terpene variation.
To resolve issues of poor reproducibility of terpene profiles in cannabis, it will be essential to perform
rigorous studies with a diversity of cannabis genotypes grown under controlled environmental
conditions and analyze terpene profiles quantitatively and qualitatively over the course of plant
development. This would need to include organ-, tissue- and cell-type specific terpene analysis, and
would have to include controlled experiments to assess effects of environmental conditions such as
light, irrigation, and nutrients. Such experiments should include not only terpene metabolite analysis,
but also a comprehensive transcriptome profiling of CsTPS gene expression. The results of such a study
would enable much needed proper assignment of reproducible terpene profiles to different strains and
support the standardization of cannabis varieties and derived consumer products.
3. Biosynthesis of cannabinoids
Compared to terpene biosynthesis, cannabinoid biosynthesis has been a priority of the limited research
on metabolite biosynthesis in cannabis to date. Much of the core cannabinoid biosynthetic pathway has
been characterized [14, 15, 16, 17]. The primary branch-point intermediate for cannabinoid biosynthesis
is cannabigerolic acid (CBGA). CBGA is produced by the prenylation of the aromatic olivetolic acid with a
geranyl moiety. An aromatic prenyltransferase (aPT) was recently cloned and shown to be active in a
metabolically engineered yeast to produce CBGA [16], and a related cannabis membrane protein with
prenyltransferase activity was previously reported in the patent literature [18]. Similar enzymes were
shown to prenylate acylphloroglucinols to produce bitter acids in hop, a close relative of cannabis [19,
20]. The precise origin of the fatty acid precursors of olivetolic acid is unknown. Genes of three different
cannabinoid synthases THCA synthase (THCAS), CBDA synthase (CBDAS), and cannabichromenic acid
synthase (CBCAS), have been published[21, 22, 23]. However, the genes and enzymes responsible for
the many minor cannabinoids, including propyl sidechain variants, remain unknown.
4. Effects attributed to terpenes in cannabis
Arguably, the only effect of cannabis terpenes on humans that is unquestionable are the fragrance
attributes of different mono- and sesquiterpene volatiles and their mixtures. Depending on the variable
composition of cannabis terpene profiles, different strains elicit different fragrance impressions, which
may affect consumer preference [24]. However, other attributes assigned to terpenes in cannabis
products, including medicinal properties, remain for now outside of the space of scientific evidence.
The so-called ‘entourage effect’ is a popular idea. It suggests a pharmacological synergy between
cannabinoids and other components of cannabis resin, in particular terpenes [25, 26]. Putative aspects
of the entourage effect include the treatment of depression, anxiety, addiction, epilepsy, cancer, and
infections. The anecdotal notion of a synergistic effect appears to stem from the perception among
cannabis users that different strains have different physiological effects. There is no doubt that the
large chemical space of thousands of plant terpenes and terpenoids includes many biologically active
molecules. Some terpenoids, such as the anticancer drug taxol, are potent and highly valuable
pharmaceuticals, the effects of which are supported by the full range of pharmacological and clinical
studies. In one of the few examples of the entourage effect being tested, terpenes were found not to
contribute to cannabinoid-mediated analgesia in rats [27]. With the possible exception of the
sesquiterpene β-caryophyllene, no molecular mechanism has been demonstrated to explain a potential
synergy of terpenes with cannabinoids. One potential explanation for the effects attributed to terpenes
is revealed in a recent review [28], pointing out that the placebo effect is partially mediated through the
endocannabinoid system, which may explain some of the perceived effects of cannabis products.
The sesquiterpene β-caryophyllene is prominent in many cannabis strains and products. The molecule
binds to the mammalian CB2 cannabinoid receptor, which may provide a plausible mechanism for
interaction with cannabinoids and a starting point for future research [29]. β-caryophyllene is one of the
least variable terpene components of cannabis (Table 1), which would suggest that it cannot explain
strain-specific effects in humans. The proposed synergistic effects of terpenes in the effects of cannabis
in humans is an area that will require careful research, which will now be possible in those jurisdictions
in which some of the legal restrictions have been lifted.
5. Claims of anticancer effects of cannabis and cannabis terpenes may do more harm than good
Certain monoterpenes have been shown to block tumor formation or inhibit cell cycle progression in
vivo and in rats [30, 31, 32]. However, the amounts of terpenes required to produce anti-proliferative
effects in rats are excessively high with up to 10% of the animals’ diet [30]. Similarly, cannabinoids may
inhibit tumor formation in animal models of cancer [33]. Laboratory studies such as these may have led
to the suggestion that cannabis extracts, with their combination of cannabinoids and terpenes, have
anti-cancer properties [25, 26]. However, to our knowledge, there is no conclusive evidence to support
claims of anticancer activity of terpenes consumed with cannabis products. While the ethanolic extract
of cannabis flowers has higher antitumor activity than pure THC, this effect was not attributed to any of
the five most abundant terpenes [35].
In general, it is important to remember that cannabis is often consumed by smoking or as a vapor. This
includes cannabis consumption by young adults. Consumer habits such as inhaling combusted or
vaporized cannabis products must be considered a health risk, including the potential risk of causing
cancer or other health issues [35, 36], before promoting unsupported claims of anti-cancer effects of
6. Perspective and future directions
Genomics has been slow to reach cannabis, largely due to legal restrictions on funding agencies and
researchers. A first reference quality cannabis genome was published in 2018 [23], enabling the
genome-wide analysis of genes for metabolic pathways systems in cannabis. More genotyping and
sequencing studies are required to encompass the full diversity of the species. A special emphasis is
needed on Eurasian and African landraces, which have been under-sampled. Critical tools for functional
genomics of metabolic systems, and ultimately crop improvement, such as genetic transformation or
genome editing, are not yet established for cannabis research in the public domain. Beyond the genes
that encode enzymes for the biosynthesis of terpenes and cannabinoids in cannabis, research is needed
to elucidate the factors that control expression of these biosynthetic systems. This would include, for
example, the regulation of cell-type specific gene expression in the context of the development of
glandular trichomes, plant architecture, and onset of female flowering.
As restrictions on research with cannabis relax, cannabis is likely to become a more popular research
organism both for the gain of basic and applied knowledge. Cannabis is a useful system for terpene
research as it produces a large volume of a diverse terpene-rich resin on its trichome-covered surfaces.
The abundance and size of its glandular trichomes make it a useful system for research in cell
specialization and regulation of terpene and cannabinoid metabolism.
At present, of the hundreds of terpene and cannabinoid metabolites that have been identified in
cannabis, the biosynthesis of less than 30 has been characterized. Future biochemical and functional
work on biosynthetic systems in cannabis would benefit from a focused community effort to produce
and archive a complete and reproducible set of metabolite and genomic data for one or a few genotypes
that will serve as a reference framework. In parallel, a larger number of cannabis types need to be
properly genotyped and phenotypically characterized (e.g. with regard to their metabolites) to
overcome current issues with inconsistencies in what is referred to as strains. The goal would be to
establish reproducible cannabis varieties for use in research and in the industry, comparable to the well-
defined grapevine varieties that are used in viticulture. Moving from strains to varieties will require the
cooperation of cannabis researchers, breeders and growers. To our knowledge, so far, no industry
association has taken a lead to set community standards and practices or define community-accessible
varieties. Researchers and industry in Canada, as the first developed nation to have fully legalized
cannabis, are uniquely positioned to lead this effort.
We thank Samuel Livingston for contributions to Figure 1. We thank Sharon Jancsik, Angela Chiang, and
Dr. Sandra Irmisch for comments on the manuscript. Research on cannabis terpenes in the Bohlmann
lab is supported with funds from Genome British Columbia (SIP001 to JB) and the Natural Sciences and
Engineering Research Council of Canada (NSERC). JKB is the recipient of an NSERC graduate student
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Figure Legends
Figure 1. Cannabis inflorescence and stalked glandular trichomes. A) Apical inflorescence from the
strain Purple Kush, eight weeks post onset of flowering. B) Floret cluster from the strain Lemon Skunk,
five weeks post onset of flowering. C) Stalked glandular trichomes on the surface of strain Finola
pistillate flowers. Scanning electron microscopy and image credit for C) thanks to Samuel Livingston,
UBC, Department of Botany.
Figure 2. Schematic of terpene and cannabinoid biosynthesis in cannabis. 5-Carbon isoprenoid building
blocks isopententyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP) are condensed to form
geranyl diphosphate (GPP) (C10) or farnesyl diphosphate (FPP) (C15). Terpene synthases (TPS) convert
GPP or FPP into terpenes. Aromatic prenyltransferases (aPTs) condense GPP with olivetolic acid to form
cannabigerolic acid (CBGA), which is cyclized by cannabinoid synthases to produce cannabinoids.
Cannabinoids: C1: cannabigerolic acid, C2: cannabichromenic acid, C3: cannabidiolic acid, C4:
tetrahydrocannabinolic acid. Monoterpenes: M1: β-pinene, M2: α-pinene, M3: β-thujone, M4: 3-carene,
M5: terpinolene, M6: limonene, M7: terpineol, M8: 1,8-cineole, M9: α-terpinene, M10: linalool, M11:
myrcene, M12: (Z)-β-ocimene. Sesquiterpenes S1: α-elemol, S2: (E)-β-farnesol, S3: (E)-β-farnesene, S4:
bisabolol, S5: (+)-α-bergamotene, S6: δ-cadinene, S7: γ-eudesmol, S8: valencene, S9: eremophilene, S10:
β-himachalene, S11: α-guaiene, S12: germacrene D, S13: alloaromadendrene, S14: β-caryophyllene.
Table 1. Publications listing cannabis terpene profiles. Purpose refers to the stated objective of the study.
Origin of plant material indicates what the authors stated as the source of their cannabis or extracts. Number
of terpenes identified includes all named or numbered compounds listed by the authors, including those not
identified using authentic standards. Publications are listed in order of date published, from earliest to most
# of terpenes
Origin of plant
Purpose of analysis
Wild-grown in
Plant Biology
Forensic samples
Grown by
Plant Biology
researchers, law
Grown by
Plant Biology
Bedrocan BV
Grown by
Metabolite survey
Grown by
Metabolite survey
Coffee shops in the
Netherlands and
Bedrocan BV
Bedrocan BV
Forensic samples
Metabolite survey
Grown outdoors
Indoor cultivator in
Submissions from
medical patients
Grown by
Plant Biology
Bedrocan BV
Bedrocan BV
Submitted by
Licensed producers
in Canada
Indoor cultivator in
New Mexico,
assorted growers
Dispensary in
Grown outdoors
... To validate that we observe patterns expected from previous studies, we first looked for correlations between specific terpene pairs. We chose pairs that have been previously observed to display robust positive correlations, likely stemming from constraints on their biochemical synthesis [63][64][65]. Strong positive correlations were seen between α-and β-pinene (Fig 3B; r To systematically understand relationships between all terpene pairs, we performed hierarchical clustering on all pairwise correlations among terpenes (Fig 4A; see Methods). ...
... We also plotted this data in the form of a network diagram configured to display connections between terpenes with the strongest correlations ( Fig 4B). This diagram provides a more compact picture of terpene co-occurrence and likely reflects the underlying biosynthesis pathways that give rise to these correlations [63][64][65]. ...
... constraints of terpene biosynthesis [63,64,69], as has been observed elsewhere [50,65]. The pooled dataset also displays features seen in sample populations from US states not represented here [35]. ...
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The legal status of Cannabis is changing, fueling an increasing diversity of Cannabis -derived products. Because Cannabis contains dozens of chemical compounds with potential psychoactive or medicinal effects, understanding this phytochemical diversity is crucial. The legal Cannabis industry heavily markets products to consumers based on widely used labeling systems purported to predict the effects of different “strains.” We analyzed the cannabinoid and terpene content of commercial Cannabis samples across six US states, finding distinct chemical phenotypes (chemotypes) which are reliably present. By comparing the observed phytochemical diversity to the commercial labels commonly attached to Cannabis -derived product samples, we show that commercial labels do not consistently align with the observed chemical diversity. However, certain labels do show a biased association with specific chemotypes. These results have implications for the classification of commercial Cannabis , design of animal and human research, and regulation of consumer marketing—areas which today are often divorced from the chemical reality of the Cannabis -derived material they wish to represent.
... In C. sativa, the TPS family is highly diverse, comprising at least 55 different members but subtle evolutionary changes to just a few amino acids of an existing TPS have resulted in altered product profiles (Allen et al., 2019). Similarly, the terpene profile of C. sativa is also highly variable, fluctuating across chemovars and averaging 11 terpenes at more than 1% total terpene content (Allen et al., 2019;Booth and Bohlmann, 2019). Analysis of 240 cannabis cultivars showed that the most abundant terpenes were β-myrcene, β-caryophyllene, D-limonene, β-pinene and α-pinene (Allen et al., 2019). ...
... Recombinant production of C. sativa enzymes has been performed using a limited range of expression systems (Booth et al., 2017Günnewich et al., 2007;Livingston et al., 2020;Zager et al., 2019). Heterologous expression of C. sativa TPSs has exclusively been performed in E. coli (Booth and Bohlmann, 2019). A combination of synthetic and trichome extracted RNA has been used to generate TPS cDNAs for transformation into expression vectors. ...
Full-text available
Cannabis sativa L. has been known for at least 2000 years as a source of important, medically significant specialised metabolites and several bio-active molecules have been enriched from multiple chemotypes. However, due to the many levels of complexity in both the commercial cultivation of cannabis and extraction of its specialised metabolites, several heterologous production approaches are being pursued in parallel. In this review, we outline the recent achievements in engineering strategies used for heterologous production of cannabinoids, terpenes and flavonoids along with their strength and weakness. We provide an overview of the specialised metabolism pathway in C. sativa and a comprehensive list of the specialised metabolites produced along with their medicinal significance. We highlight cannabinoid-like molecules produced by other species. We discuss the key biosynthetic enzymes and their heterologous production using various hosts such as microbial and eukaryotic systems. A brief discussion on complementary production strategies using co-culturing and cell-free systems is described. Various approaches to optimise specialised metabolite production through co-expression, enzyme engineering and pathway engineering are discussed. We derive insights from recent advances in metabolic engineering of hosts with improved precursor supply and suggest their application for the production of C. sativa speciality metabolites. We present a collation of non-conventional hosts with speciality traits that can improve the feasibility of commercial heterologous production of cannabis-based specialised metabolites. We provide a perspective of emerging research in synthetic biology, allied analytical techniques and plant heterologous platforms as focus areas for heterologous production of cannabis specialised metabolites in the future.
... 27 Cannabis odors are largely attributable to a class of volatile organic compounds (VOCs) called terpenes. 28 More than 150 distinct terpenes have been identified in different cannabis strains, 29 from among approximately 30,000 expressed across the plant world. 30 In their review, the authors identified a few key terpenes often detected at high levels in and around grows: myrcene (earthy, A few key terpenes are often detected at high levels in and around cannabis grows, yielding scents that may be piney, grassy, citrusy, woody, floral, or earthy. ...
... In addition to phytocannabinoids, the Cannabis genus also produces non-cannabinoid compounds, including terpenoids (12), which may also be medically useful (13)(14)(15). The pharmacological contributions of minor cannabinoids and non-cannabinoid compounds have been highlighted and popularized under the term "entourage effect" (16,17). ...
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Cannabis is one of the few plant genera capable of producing cannabinoids, the effects of which are synergized by terpene interactions. As the biosynthesis of both metabolite classes requires the same intracellular feedstocks, this work describes the coordinated regulation of global metabolic pathways that allows for their joint copious production in vivo. To this end, a transcriptomics-based approach to characterize the glandular trichomes of five Cannabis cultivars was pursued. Besides revealing metabolic traits that enhanced and proportionated the supply of critical carbon precursors, in-depth analysis showed significantly increased gene expression of two particular enzymes to meet the huge nicotinamide adenine dinucleotide phosphate (NADPH) demand of secondary metabolite production. Furthermore, it led to a hypothesis that the methyl-d-erythritol 4-phosphate pathway might be utilized more than the mevalonic acid pathway in Cannabis trichomes. While both pathways were found to be activated in a modular and calibrated way that reflected their broad participation in physiological processes, the genes for hexanoate, cannabinoid, and terpene biosynthesis were, in contrast, up-regulated in an en bloc and multi-loci manner due to their specific roles in secondary metabolite production. In addition, three new terpene synthases were characterized based on both in silico and experimental assays. Altogether, the study enhances the current understanding of secondary metabolite production in Cannabis cultivars, which may assist in their characterization and development.
Cannabis is the most versatile species. Hemp and marijuana have been used for fibre, oil, medicinal and recreational purposes from millennia. Throughout the last century, the plant has been generally outlawed because of its psychotropic effects in many nations. In recent past, the studies on cannabis revealed the evidence of its high medicinal properties and its uses in treating life threatening diseases, which leads to the relaxation of legislation in many counties. Now, the genetic and genomics as well as the cannabis derived products enjoys renewed attention. In this chapter, the discussion was made on the advent of genomics and breeding strategies to improve various traits of cannabis. This will bring insights on future direction of cannabis breeding.
Thirdhand smoke (THS) describes the persistent contamination on indoor surfaces following smoking. Cannabis THS is a chemically distinct form of THS which remains poorly characterized, relative to tobacco. Understanding its fate is necessary to mitigate exposure to cannabis smoke components and potentially harmful transformation products. In this work, the heterogeneous reaction of surface-bound Δ9-tetrahydrocannabinol (THC), a component of cannabis smoke, with ozone was examined. Cannabis smoke deposits were collected inside a Teflon chamber on glass, cotton, and tile surfaces. Samples were exposed to high (100 ppb) and realistic indoor ozone concentrations (20 ppb) inside a flow tube and to low ozone levels (2 to 29 ppb) present in a genuine indoor environment. Chemical transformations were monitored by liquid chromatography-tandem mass spectrometry (LC-MS/MS), with epoxide and dicarbonyl products detected. Rapid initial loss of THC was observed on all surfaces, but a small fraction (5%-10%) persisted for extended time scales. A short loss lifetime (a few hours) was observed for THC deposited on glass and tile under typical indoor ozone exposures, leading to almost complete loss at longer times. THC decay on cotton was approximately five times longer than on glass, with up to 10% of THC remaining after 1 week of exposure.
The extraction of compounds present in Cannabis sativa biomass from Colombian crops was studied using a semi-continuous lixiviation process. To this effect, three extraction stages were implemented, in which successive transfers were carried out at fixed times, seeking to emulate a continuous countercurrent process. In this way, the intention is to provide the Colombian agroindustry with an extraction method that is efficient and requires a lower initial investment than other techniques such as supercritical fluid extraction. Absolute ethanol was used as solvent, and constant temperature and stirring speed were applied. The obtained results indicate that, once the process is stabilized, extracted mass percentages (with respect to the total mass on a dry basis) of 10,5% at 40 °C and 9,5% at 19 °C are achieved, which are competitive compared to the 11,07% achieved through Soxhlet extraction. Besides, the proposed process is clearly a better alternative than a single-stage extraction method, through which it was possible to extract 5% of the mass. Five cannabinoids were identified in the obtained extract, and, with the proposed process, it was possible to extract 66% of cannabidiol present in the original biomass.
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There has been an increased use of medical Cannabis in the United States of America as more states legalize its use. Complete chemical analyses of this material can vary considerably between producers and is often not fully provided to consumers. As phytochemists in a state with legal medical Cannabis we sought to characterize the accumulation of phytochemicals in material grown by licensed commercial producers. We report the development of a simple extraction and analysis method, amenable to use by commercial laboratories for the detection and quantification of both cannabinoids and terpenoids. Through analysis of developing flowers on plants, we can identify sources of variability of floral metabolites due to flower maturity and position on the plant. The terpenoid composition varied by accession and was used to cluster cannabis strains into specific types. Inclusion of terpenoids with cannabinoids in the analysis of medical cannabis should be encouraged, as both of these classes of compounds could play a role in the beneficial medical effects of different cannabis strains.
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The smell of marijuana (Cannabis sativa L.) is of interest to users, growers, plant breeders, law enforcement and, increasingly, to state-licensed retail businesses. The numerous varieties and strains of Cannabis produce strikingly different scents but to date there have been few, if any, attempts to quantify these olfactory profiles directly. Using standard sensory evaluation techniques with untrained consumers we have validated a preliminary olfactory lexicon for dried cannabis flower, and characterized the aroma profile of eleven strains sold in the legal recreational market in Colorado. We show that consumers perceive differences among strains, that the strains form distinct clusters based on odor similarity, and that strain aroma profiles are linked to perceptions of potency, price, and smoking interest.
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Inhalable, noncombustible cannabis products are playing a central role in the expansion of the medical and recreational use of cannabis. In particular, the practice of “dabbing” with butane hash oil has emerged with great popularity in states that have legalized cannabis. Despite their growing popularity, the degradation product profiles of these new products have not been extensively investigated. The study herein focuses on the chemistry of myrcene and other common terpenes found in cannabis extracts. Methacrolein, benzene, and several other products of concern to human health were formed under the conditions that simulated real-world dabbing. The terpene degradation products observed are consistent with those reported in the atmospheric chemistry literature.
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Cannabis (Cannabis sativa) plants produce and accumulate a terpene-rich resin in glandular trichomes, which are abundant on the surface of the female inflorescence. Bouquets of different monoterpenes and sesquiterpenes are important components of cannabis resin as they define some of the unique organoleptic properties and may also influence medicinal qualities of different cannabis strains and varieties. Transcriptome analysis of trichomes of the cannabis hemp variety 'Finola' revealed sequences of all stages of terpene biosynthesis. Nine cannabis terpene synthases (CsTPS) were identified in subfamilies TPS-a and TPS-b. Functional characterization identified mono-and sesqui-TPS, whose products collectively comprise most of the terpenes of 'Finola' resin, including major compounds such as β-myr-cene, (E)-β-ocimene, (-)-limonene, (+)-α-pinene, β-caryophyllene, and α-humulene. Transcripts associated with terpene biosynthesis are highly expressed in trichomes compared to non-resin producing tissues. Knowledge of the CsTPS gene family may offer opportunities for selection and improvement of terpene profiles of interest in different cannabis strains and varieties.
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Introduction: With laws changing around the world regarding the legal status of Cannabis sativa (cannabis) it is important to develop objective classification systems that help explain the chemical variation found among various cultivars. Currently cannabis cultivars are named using obscure and inconsistent nomenclature. Terpenoids, responsible for the aroma of cannabis, are a useful group of compounds for distinguishing cannabis cultivars with similar cannabinoid content. Methods: In this study we analyzed terpenoid content of cannabis samples obtained from a single medical cannabis dispensary in California over the course of a year. Terpenoids were quantified by gas chromatography with flame ionization detection and peak identification was confirmed with gas chromatography mass spectrometry. Quantitative data from 16 major terpenoids were analyzed using hierarchical clustering analysis (HCA), principal component analysis (PCA), partial least squares discriminant analysis (PLS-DA), and orthogonal partial least squares discriminant analysis (OPLS-DA). Results: A total of 233 samples representing 30 cultivars were used to develop a classification scheme based on quantitative data, HCA, PCA, and OPLS-DA. Initially cultivars were divided into five major groups, which were subdivided into 13 classes based on differences in terpenoid profile. Different classification models were compared with PLS-DA and found to perform best when many representative samples of a particular class were included. Conclusion: A hierarchy of terpenoid chemotypes was observed in the data set. Some cultivars fit into distinct chemotypes, whereas others seemed to represent a continuum of chemotypes. This study has demonstrated an approach to classifying cannabis cultivars based on terpenoid profile.
Cannabis sativa is widely cultivated for medicinal, food, industrial, and recreational use, but much remains unknown regarding its genetics, including the molecular determinants of cannabinoid content. Here, we describe a combined physical and genetic map derived from a cross between the drug-type strain Purple Kush and the hemp variety “Finola.” The map reveals that cannabinoid biosynthesis genes are generally unlinked but that aromatic prenyltransferase (AP), which produces the substrate for THCA and CBDA synthases (THCAS and CBDAS), is tightly linked to a known marker for total cannabinoid content. We further identify the gene encoding CBCA synthase (CBCAS) and characterize its catalytic activity, providing insight into how cannabinoid diversity arises in cannabis. THCAS and CBDAS (which determine the drug vs. hemp chemotype) are contained within large (>250 kb) retrotransposon-rich regions that are highly nonhomologous between drug- and hemp-type alleles and are furthermore embedded within -40 Mb of minimally recombining repetitive DNA. The chromosome structures are similar to those in grains such as wheat, with recombination focused in gene-rich, repeat-depleted regions near chromosome ends. The physical and genetic map should facilitate further dissection of genetic and molecular mechanisms in this commercially and medically important plant.
Cannabaceae plants Cannabis sativa L. and Humulus lupulus L. are rich in terpenes-both are typically comprised of terpenes as up to 3-5% of the dry-mass of the female inflorescence. Terpenes of cannabis and hops are typically simple mono-and sesquiterpenes derived from two and three isoprene units, respectively. Some terpenes are relatively well known for their potential in biomedicine and have been used in traditional medicine for centuries, while others are yet to be studied in detail. The current, comprehensive review presents terpenes found in cannabis and hops. Terpenes' medicinal properties are supported by numerous in vitro, animal and clinical trials and show anti-inflammatory, antioxidant, analgesic, anticonvulsive, antidepressant, anxiolytic, anticancer, antitumor, neuroprotective, anti-mutagenic, anti-allergic, antibiotic and anti-diabetic attributes, among others. Because of the very low toxicity, these terpenes are already widely used as food additives and in cosmetic products. Thus, they have been proven safe and well-tolerated.
Breast cancer is the second leading cause of death among women. Although early diagnosis and development of new treatments have improved their prognosis, many patients present innate or acquired resistance to current therapies. New therapeutic approaches are therefore warranted for the management of this disease. Extensive preclinical research has demonstrated that cannabinoids, the active ingredients of Cannabis sativa, trigger antitumor responses in different models of cancer. Most of these studies have been conducted with pure compounds, mainly Δ9-tetrahydrocannabinol (THC). The cannabis plant, however, produces hundreds of other compounds with their own therapeutic potential and the capability to induce synergic responses when combined, the so-called "entourage effect". Here, we compared the antitumor efficacy of pure THC with that of a botanical drug preparation (BDP). The BDP was more potent than pure THC in producing antitumor responses in cell culture and animal models of ER+/PR+, HER2+ and triple-negative breast cancer. This increased potency was not due to the presence of the 5 most abundant terpenes in the preparation. While pure THC acted by activating cannabinoid CB2 receptors and generating reactive oxygen species, the BDP modulated different targets and mechanisms of action. The combination of cannabinoids with estrogen receptor- or HER2-targeted therapies (tamoxifen and lapatinib, respectively) or with cisplatin, produced additive antiproliferative responses in cell cultures. Combinations of these treatments in vivo showed no interactions, either positive or negative. Together, our results suggest that standardized cannabis drug preparations, rather than pure cannabinoids, could be considered as part of the therapeutic armamentarium to manage breast cancer.