Cannabis Chemovars Research Articles

Identification of 69 magnesium-free chlorophyll derivatives in decarboxylated extracts of medical Cannabis chemovars using ESI-LC-HRMS/MS

The Cannabis plant contains many groups of bioactive metabolites with potential pharmacological properties. These properties have mainly been attributed to cannabinoids and terpenoids, with other groups receiving little attention. The usefulness of the abundant group of chlorophyll derivatives (CDs) was illustrated in various applications, but little is known regarding their presence and significance in Cannabis. We hypothesized that the heating accompanying the process extract preparation would result in a pool of CDs that have lost the central magnesium (Mg) ion. Herein, we introduce a liquid chromatography-tandem mass spectrometry approach for separating, identifying, and quantifying Mg-free CDs in Cannabis extracts after decarboxylation. We assessed 14 Cannabis cultivars representing the four types of chemovars and identified 69 distinct Mg-free CDs. These were separated into five families: Pheophorbide a, Pheophorbide b, Pheophytin a, Pheophytin b, and Purpurin 18. The structure of the Mg-free CDs was determined through their MS/MS fragmentation spectra as phytylated derivatives compared to dephytylated ones, and then further to pyroderivatives, alkylated and oxidized CDs. Substantial variation was found between the four different chemovar types and within different cultivars of the same chemovar, with the family of Pheophytin a, the most prevalent in all extracts. Type III chemovar high-cannabidiol plants contained significantly higher amounts of Mg-free CDs. These differences in abundance may result in variations in the therapeutic effects of plants that are considered similar according to their cannabinoid profiles.

Comparative chemometric modeling of fresh and dry cannabis inflorescences using FT-NIR spectroscopy: Quantification and classification insights.

Cannabis sativa L. inflorescences are rich in cannabinoids and terpenes. Traditional chemical analysis methods for cannabinoids and terpenes, such as liquid and gas chromatography (using UV or MS detectors), are expensive and time-consuming. This study explores the use of Fourier transform near-infrared (FT-NIR) spectroscopy combined with chemometric approaches for classifying cannabis chemovars and predicting cannabinoid and terpene concentrations for the first time in freshly harvested (wet) cannabis inflorescence. The study also compares the performance of FT-NIR spectroscopy on wet versus dry cannabis inflorescences. Spectral data from 187 samples across seven cannabis chemovars were analyzed using partial least squares-discriminant analysis (PLS-DA) and partial least squares-regression (PLS-R) models. The PLS-DA models effectively classified chemovars and major classes using only two latent variables (LVs) with minimal overfitting risk, with sensitivity, specificity, and accuracy values approaching 1. Despite the high water content in wet cannabis inflorescence, the PLS-R models demonstrated good to excellent predictive capabilities for nine cannabinoids and eight terpenes using FT-NIR spectra for the first time, achieving cross-validation and prediction R-squared values greater than 0.7, ratio of performance to interquartile range (RPIQ) exceeding 2, and a RMSECV/RMSEC ratio below 1.24. However, the low-cannabidiolic acid submodel and (-)-Δ9-trans-tetrahydrocannabinol model showed poor predictive performance. Some cannabinoid and terpene prediction models in wet cannabis inflorescence exhibited lower predictive capabilities compared with previously published models for dry cannabis inflorescence. These findings suggest that FT-NIR spectroscopy can be a viable rapid on-site analytical tool for growers during the inflorescence flowering stage.

Sink strength, nutrient allocation, cannabinoid yield and associated transcript profiles vary in two drug-type Cannabis chemovars.

Cannabis sativa L. is one of the oldest domesticated crops. Hemp-type cultivars, which predominantly produce non-intoxicating cannabidiol (CBD), have been selected for their fast growth, seed, and fibre production, while drug-type chemovars were bred for high accumulation of tetrahydrocannabinol (THC). We investigated how the generation of CBD-dominant chemovars by introgression of hemp- into drug-type Cannabis impacted plant performance. The THC-dominant chemovar showed superior sink strength, higher flower biomass and demand-driven control of nutrient uptake. By contrast, the CBD-dominant chemovar hyperaccumulated phosphate in sink organs leading to reduced carbon and nitrogen assimilation in leaves, which limited flower biomass and cannabinoid yield. RNA-seq analyses determined organ- and chemovar-specific differences in expression of genes associated with nitrate and phosphate homeostasis as well as growth-regulating transcription factors that were correlated with measured traits. Among these were genes positively selected for during Cannabis domestication encoding an inhibitor of the phosphate starvation response SPX DOMAIN GENE3, nitrate reductase and two nitrate transporters. Altered nutrient sensing, acquisition or distribution are likely a consequence of adaption to growth on marginal, low-nutrient input lands in hemp. Our data provide evidence that such ancestral traits may become detrimental for female flower development and consequently overall CBD yield in protected cropping environments.

Metabolic characterization and transcriptional profiling of polyphenols in Cannabis sativa L. inflorescences with different chemical phenotypes.

After the most comprehensive analysis of the phenolic composition in Cannabis reported to date, a total of 211 compounds were identified, phenolic profiles were able to discriminate cannabis varieties and a complex regulatory network for phenolics accumulation in Cannabis chemovars was highlighted. Female inflorescences of Cannabis sativa L. are plenty of secondary metabolites, of which flavonoids and phenolic acids have been investigated by far less than phytocannabinoids and terpenoids. Understanding the biochemical composition in phenylpropanoids of Cannabis inflorescences, the molecular basis of flavonoid synthesis and how their content can be modulated by specific transcription factors will shed light on the variability of this trait in the germplasm, allowing the identification of biologically active metabolites that can be of interest to diverse industries. In this work, an untargeted metabolomic approach via UHPLC-HRMS was adopted to investigate the composition and variability of phenylpropanoids in thirteen Cannabis genotypes differentiated for their profile in phytocannabinoids, highlighting that phenolic profiles can discriminate varieties, with characteristic, unique genotype-related patterns. Moreover, the transcription profile of candidate phenolics regulatory MYB and bHLH transcription factors, analyzed by RT-qPCR, appeared strongly genotype-related, and specific patterns were found to be correlated between biochemical and transcriptional levels. Results highlight a complex regulatory network for phenolic accumulation in Cannabis chemovars that will need further insights from the functional side.

Untargeted cannabinomics reveals the chemical differentiation of industrial hemp based on the cultivar and the geographical field location

Cannabis sativa has long been harvested for industrial applications related to its fibers. Industrial hemp cultivars, a botanical class of Cannabis sativa with a low expression of intoxicating Δ9-tetrahydrocannabinol (Δ9-THC) have been selected for these purposes and scarcely investigated in terms of their content in bioactive compounds. Following the global relaxation in the market of industrial hemp-derived products, research in industrial hemp for pharmaceutical and nutraceutical purposes has surged. In this context, metabolomics-based approaches have proven to fulfill the aim of obtaining comprehensive information on the phytocompound profile of cannabis samples, going beyond the targeted evaluation of the major phytocannabinoids. In the present paper, an HRMS-based metabolomics study was addressed to seven distinct industrial hemp cultivars grown in four experimental fields in Northern, Southern, and Insular Italy. Since the role of minor phytocannabinoids as well as other phytocompounds was found to be critical in discriminating cannabis chemovars and in determining its biological activities, a comprehensive characterization of phytocannabinoids, flavonoids, and phenolic acids was carried out by LC-HRMS and a dedicated data processing workflow following the guidelines of the metabolomics Quality Assurance and Quality Control Consortium. A total of 54 phytocannabinoids, 134 flavonoids, and 77 phenolic acids were annotated, and their role in distinguishing hemp samples based on the geographical field location and cultivar was evaluated by ANOVA-simultaneous component analysis. Finally, a low-level fused model demonstrated the key role of untargeted cannabinomics extended to lesser-studied phytocompound classes for the discrimination of hemp samples.

Development and Validation of a GC-FID Method for the Quantitation of 20 Different Acidic and Neutral Cannabinoids.

For decades, Cannabis sativa had been illegal to sell or consume around the world, including in the United States. However, in light of the recent 2018 Farm Bill and the legalization of hemp across the US, various cannabis preparations have flooded the market, making it essential to be able to quantitate the levels of the different acidic and neutral cannabinoids in C. sativa and to have a complete cannabinoid profile of the different chemovars of the cannabis plant. A GC-FID method was developed and validated for the analysis of 20 acidic and neutral cannabinoids as trimethylsilyl (TMS) derivatives. The analyzed cannabinoids include cannabidivarinic acid (CBDVA), cannabidiolic acid (CBDA), cannabinolic acid (CBNA), cannabielsoic acid (CBEA), cannabicyclolic acid (CBLA), cannabichromenic acid (CBCA), trans-Δ9-tetrahydrocannabivarinic acid (Δ9-THCVA), trans-Δ9-tetrahydrocannabinolic acid A (Δ9-THCAA), cannabigerolic acid (CBGA), cannabidiol (CBD), cannabicyclol (CBL), cannabidivarin (CBDV), trans-Δ9-tetrahydrocannabivarin (THCV), cannabichromene (CBC), trans-Δ8-tetrahydrocannabinol (Δ8-THC), trans-Δ9-tetrahydrocannabinol (Δ9-THC), cannabigerol (CBG), cannabinol (CBN), cannabicitran (CBT), and cannabielsoin (CBE). The method limit of detection (LOD) was as low as 0.1 µg/mL, while the limit of quantitation ranged from 0.25 µg/mL to 0.5 µg/mL. The precision (%RSD) was < 10%, while trueness ranged from 90 - 107%. The developed method is simple, accurate, and sensitive for the quantitation of all 20 acidic and neutral cannabinoids. Finally, the proposed method was successfully applied to the quantitation of the cannabinoids in different cannabis chemovars grown at the University of Mississippi.

Acute effects of cannabinoid consumption on endocannabinoid levels

Investigating the acute effects of cannabinoids consumption on circulating levels of endocannabinoids. Cannabis remains the most commonly used drug of abuse worldwide and the legalization in several states caused a drastic change in the variety of products available on the market. Over the past few years, several studies reported that the average concentration of Δ9-tetrahydrocannabinol (THC) drastically increased as well as the availability of products containing different combinations of THC and other cannabinoids, mainly cannabidiol (CBD). However, little is known about the acute effects of THC and/or CBD on the lipidome and more specifically the endocannabinoid system. The aim of the present study was to examine the acute effects of three cannabis flower chemovars with different THC to CBD ratios, in order to test whether chemovars with a higher CBD content produce different subjective effects and analyze their influence on the circulating levels of endocannabinoids. Participants were randomly assigned to ad libitum administration of one of three chemovars (THC-dominant: 24% THC, 1% CBD; THC + CBD: 9% THC, 10% CBD; CBD-dominant: 1% THC, 23% CBD); plasma samples were collected in a mobile pharmacology laboratory before, immediately after and 1 hour after ad libitum administration of their assigned chemovar. Twelve cannabinoids and 14 endocannabinoids were quantified using liquid chromatography tandem mass spectrometry assays previously published and validated by our group. Statistical analysis was performed using SPSS and MetaboAnalyst. One hundred and six participants were included in the study; 29 were assigned to the THC-dominant chemovar, 37 to the CBD-dominant chemovar and 40 to the THC + CBD chemovar. A total of 318 plasma samples were analyzed for both cannabinoids and endocannabinoids concentrations. Results were divided in 9 sub-groups based on the chemovar used and the time after consumption. Across the different time points, significant changes were noted for docosatetraenoyl ethanolamide (DEA), palmitoyl ethanolamide (PEA), stearoyl ethanolamide (SEA), linoleoyl ethanolamide (LEA) and oleoyl ethanolamine (OEA). Interestingly, anandamide (AEA) concentrations were significantly decreased only after consumption of the CBD-dominant chemovar and the THC + CBD chemovar. The mean AEA concentrations in the CBD-dominant group dropped from 0.74 ± 0.24 ng/mL before consumption to 0.54 ± 0.17 ng/mL one hour after use ( P < 0.001) and from 0.70 ± 0.18 ng/mL to 0.53 ± 0.17 ng/mL in the THC + CBD chemovar group ( P < 0.001). Chronic cannabis abuse can lead to damaging effects in humans and some authors examined whether components of the endocannabinoid system will undergo alterations and have shown decreased anandamide levels in CSF and an alteration of serum anandamide was not observed in the same study. Others reported increased levels of anandamide upon CBD treatment, mostly in Schizophrenia patients. These studies focused on long-term effects (14, 28 days). To our knowledge, this is the first study to assess the acute effects of different cannabis chemovars (THC-dominant, CBD-dominant and THC + CBD chemovar) on the circulating levels of endocannabinoids in healthy frequent user volunteers. It is very interesting that upon CBD consumption a decrease in AEA was observed. This decrease might alter the effect of THC, since AEA and THC both bind to CB1 receptors. Further studies are needed to better understand the acute effects of cannabis consumption on the circulating levels of endocannabinoids and potentially explain the differences in endocannabinoid patterns after acute versus chronic consumption as well as the potential indirect effect of CBD on THC's mechanism of action.

Multivariate classification of cannabis chemovars based on their terpene and cannabinoid profiles

Cannabis is used to treat various medical conditions, and lines are commonly classified according to their total concentrations of Δ9-tetrahydrocannabinol (THC) and cannabidiol (CBD). Based on their ratio of total THC to total CBD, cannabis cultivars are commonly classified into high-THC, high-CBD, and hybrid classes. While cultivars from the same class have similar compositions of major cannabinoids, their levels of other cannabinoids and their terpene compositions may differ substantially. Therefore, a more comprehensive and accurate classification of medicinal cannabis cultivars, based on a large number of cannabinoids and terpenes is needed. For this purpose, three different chemometric-based classification models were constructed using three sets of chemical profiles. We examined those models to determine which provides the most accurate “chemovar” classification. This was done by analyzing profiles of cannabinoids, terpenes, and the combination of these substances using the partial least square-discriminant analysis multivariate (PLS-DA) technique. The chemical profiles were selected from the three major classes of medicinal cannabis that are most commonly prescribed to patients in Israel: high-THC, high-cannabigerol (CBG), and hybrid. We studied the correlations between cannabinoids and terpenes to identify major bio-indicators representing the plant's terpene and cannabinoid content. All three PLS-DA models provided highly accurate classifications, utilizing six to nine latent variables with an overall accuracy ranging from 2 to 11% CV. The PLS-DA model applied to the combined cannabinoid-and-terpene profile did the best job of differentiating between the chemovars in terms of misclassification error, sensitivity, specificity, and accuracy. The combined cannabinoid-and-terpene PLS-DA profile had cross-validation and prediction misclassification errors of 4% and 0%, respectively. This is the first study to demonstrate the highly accurate classification of samples of medicinal cannabis based on their cannabinoid and terpene profiles, as compared to cannabinoid profiles alone. Furthermore, our correlation analysis indicated that 11 cannabinoids and terpenes might serve as bio-indicators for 32 different active compounds. These findings suggest that the use of multivariate statistics could assist in breeding studies and serve as a tool for minimizing the mislabeling of cannabis inflorescences.

Analgesic Potential of Terpenes Derived from Cannabis sativa.

Pain prevalence among adults in the United States has increased 25% over the past two decades, resulting in high health-care costs and impacts to patient quality of life. In the last 30 years, our understanding of pain circuits and (intra)cellular mechanisms has grown exponentially, but this understanding has not yet resulted in improved therapies. Options for pain management are limited. Many analgesics have poor efficacy and are accompanied by severe side effects such as addiction, resulting in a devastating opioid abuse and overdose epidemic. These problems have encouraged scientists to identify novel molecular targets and develop alternative pain therapeutics. Increasing preclinical and clinical evidence suggests that cannabis has several beneficial pharmacological activities, including pain relief. Cannabis sativa contains more than 500 chemical compounds, with two principle phytocannabinoids, Δ9-tetrahydrocannabinol (Δ9-THC) and cannabidiol (CBD). Beyond phytocannabinoids, more than 150 terpenes have been identified in different cannabis chemovars. Although the predominant cannabinoids, Δ9-THC and CBD, are thought to be the primary medicinal compounds, terpenes including the monoterpenes β-myrcene, α-pinene, limonene, and linalool, as well as the sesquiterpenes β-caryophyllene and α-humulene may contribute to many pharmacological properties of cannabis, including anti-inflammatory and antinociceptive effects. The aim of this review is to summarize our current knowledge about terpene compounds in cannabis and to analyze the available scientific evidence for a role of cannabis-derived terpenes in modern pain management. SIGNIFICANCE STATEMENT: Decades of research have improved our knowledge of cannabis polypharmacy and contributing phytochemicals, including terpenes. Reform of the legal status for cannabis possession and increased availability (medicinal and recreational) have resulted in cannabis use to combat the increasing prevalence of pain and may help to address the opioid crisis. Better understanding of the pharmacological effects of cannabis and its active components, including terpenes, may assist in identifying new therapeutic approaches and optimizing the use of cannabis and/or terpenes as analgesic agents.

A Review of the Potential Use of Pinene and Linalool as Terpene-Based Medicines for Brain Health: Discovering Novel Therapeutics in the Flavours and Fragrances of Cannabis.

“Medicinal cannabis” is defined as the use of cannabis-based products for the treatment of an illness. Investigations of cannabis compounds in psychiatric and neurological illnesses primarily focus on the major cannabinoids, cannabidiol (CBD) and Δ9-tetrahydrocannabinol (Δ9-THC), which are hypothesised to benefit multiple illnesses manifesting cognitive impairment, neurodegeneration and neuro-inflammation, as well as chronic pain, epilepsy and post-traumatic stress disorder, respectively. The cannabis plant contains >500 compounds, including terpenes responsible for the flavour and fragrance profiles of plants. Recently, research has begun providing evidence on the potential use of certain plant-derived terpenes in modern medicine, demonstrating anti-oxidant, anti-inflammatory, and neuroprotective effects of these compounds. This review examined the effects of two key terpenes, pinene and linalool, on parameters relevant to neurological and psychiatric disorders, highlighting gaps in the literature and recommendations for future research into terpene therapeutics. Overall, evidence is mostly limited to preclinical studies and well-designed clinical trials are lacking. Nevertheless, existing data suggests that pinene and linalool are relevant candidates for further investigation as novel medicines for illnesses, including stroke, ischemia, inflammatory and neuropathic pain (including migraine), cognitive impairment (relevant to Alzheimer's disease and ageing), insomnia, anxiety, and depression. Linalool and pinene influence multiple neurotransmitter, inflammatory and neurotrophic signals as well as behaviour, demonstrating psycho-activity (albeit non-intoxicating). Optimising the phytochemical profile of cannabis chemovars to yield therapeutic levels of beneficial terpenes and cannabinoids, such as linalool, pinene and CBD, could present a unique opportunity to discover novel medicines to treat psychiatric and neurological illnesses; however, further research is needed.

Phytocannabinomics: Untargeted metabolomics as a tool for cannabis chemovar differentiation

Cannabis sativa is traditionally classified according to five chemotypes based on the concentration of the main phytocannabinoids tetrahydrocannabinol (THC), cannabidiol (CBD), and cannabigerol (CBG). However, cannabis chemovars and varieties very often present similar concentrations of such phytocannabinoids but different chemical profiles, which is unavoidably translated into different pharmacological effects when used for therapeutic purposes. For this reason, a more refined approach is needed for chemovar distinction, which is described in this study and named phytocannabinomics. The classification was achieved by a comprehensive characterization of the phytocannabinoid composition, by liquid chromatography coupled to high-resolution mass spectrometry untargeted metabolomics for the detection of over a hundred phytocannabinoids, and data analysis by chemometrics for chemovars differentiation. The method was developed on fifty cannabis varieties, grown under the same conditions, and was validated to discriminate between the standard chemotypes by partial least squares discriminant analysis. Then, the method was extended to consider the entire chemical variety of the cannabis accessions, by an unsupervised approach based on the principal component analysis. The latter approach clearly indicated several new subgroups within the traditional classifications, which arise from a unique composition of the minor phytocannabinoids. The existence of these subgroups, which were never described before, is of critical importance for evaluating the pharmacological effects of cannabis chemovars.

The Cannabis Terpenes.

Terpenes are the primary constituents of essential oils and are responsible for the aroma characteristics of cannabis. Together with the cannabinoids, terpenes illustrate synergic and/or entourage effect and their interactions have only been speculated in for the last few decades. Hundreds of terpenes are identified that allude to cannabis sensory attributes, contributing largely to the consumer’s experiences and market price. They also enhance many therapeutic benefits, especially as aromatherapy. To shed light on the importance of terpenes in the cannabis industry, the purpose of this review is to morphologically describe sources of cannabis terpenes and to explain the biosynthesis and diversity of terpene profiles in different cannabis chemovars.

Cannabis Phytomolecule 'Entourage': From Domestication to Medical Use.

Cannabis has been used as a medicine for millennia. Crude extracts of cannabis inflorescence contain numerous phytomolecules, including phytocannabinoids, terpenes, and flavonoids. Combinations of phytomolecules have been recently established as superior to the use of single molecules in medical treatment owing to the 'entourage effect'. Two types of entourage effects are defined: 'intra-entourage', resulting from interactions among phytocannabinoids or terpenes, and 'inter-entourage', attributed to interactions between phytocannabinoids and terpenes. It is suggested that the phytomolecule assemblages found in cannabis chemovars today derive from selective breeding during ancient cultivation. We propose that the current cannabis chemotaxonomy should be redefined according to chemical content and medicinal activity. In parallel, combinations of phytomolecules that exhibit entourage activity should be explored further for future drug development.

A pharmacological characterization of Cannabis sativa chemovar extracts

BackgroundCannabis contains Δ9-tetrahydrocannabinol (Δ9-THC) and cannabidiol (CBD) as the primary constituents responsible for pharmacological activity. However, there are numerous additional chemically-related structures to Δ9–THC and CBD that are pharmacologically active and may influence the pharmacological properties of Δ9-THC and CBD. This study chemically characterized the cannabinoid constituents in a series of cannabis chemovar extracts and investigated the potential cannabinoid entourage effect in two behavioral assays.MethodsSix chemovar extracts were compared to pure Δ9-THC, CBD and morphine for effects on the following behavioral assays in mice: hot plate and tail suspension. The battery of behavioral tests was conducted post intravenous administration of cannabis chemovar extract. Cannabinoid profiles of extracts were analyzed using high performance liquid chromatography. Cannabis extracts were administered at equal doses of Δ9-THC to investigate the role of their cannabinoid profiles in modulating the effects of Δ9-THC. Dose response curves were fit using a log[inhibitor] vs response three parameter model and differences between group means were determined using a one-way ANOVA followed by a post hoc test.ResultsCannabis chemovars tested in this study exhibited substantially different cannabinoid profiles. All chemovars produced dose-dependent immobility in the tail suspension assay and dose-dependent antinociception in the hot plate assay. The maximum antinociceptive effect and ED50 was comparable between cannabis chemovars and Δ9-THC. Two cannabis chemovars produced significantly greater immobility in the tail suspension test, with no significant differences in ED50.ConclusionsCommercially available cannabis chemovars vary widely in cannabinoid content, but when equalized for Δ9-THC content, they produce similar behavioral effects with two exceptions. These findings provide only limited support for the entourage hypothesis. Further studies are necessary to characterize the nature of these pharmacological differences between cannabis chemovars and pure Δ9-THC.

Tandem Mass Spectrometric Quantification of 93 Terpenoids in Cannabis Using Static Headspace Injections.

The therapeutic effect of Cannabis largely depends on the content of its pharmacologically active secondary metabolites, mainly phytocannabinoids, flavonoids, and terpenoids. Recent studies suggest there are therapeutic effects of specific terpenoids as well as synergistic effects with other active compounds in the plant. Although Cannabis contains an overwhelming milieu of terpenoids, only a limited number are currently reported and used for metabolic analysis of Cannabis chemovars. In this study, we report the development and validation of a method for simultaneous quantification of 93 terpenoids in Cannabis air-dried inflorescences and extracts. This method employs the full evaporation technique via a static headspace sampler, followed by gas chromatography-mass spectrometry (SHS-GC/MS/MS). In the validation process, spiked terpenoids were quantified with acceptable repeatability, reproducibility, sensitivity, and accuracy. Three medical Cannabis chemovars were used to study the effect of sample preparation and extraction methods on terpenoid profiles. This method was further applied for studying the terpenoid profiles of 16 different chemovars acquired at different dates. Our results demonstrate that sample preparation methods may significantly impact the chemical fingerprint compared to the nontreated Cannabis. This emphasizes the importance of performing SHS extraction in order to study the natural terpenoid contents of chemovars. We also concluded that most inflorescences expressed relatively unique terpenoid profiles for the most pronounced terpenoids, even when sampled at different dates, although absolute concentrations may vary due to aging. The suggested method offers an ideal tool for terpenoid profiling of Cannabis and sets the scene for more comprehensive works in the future.

The Terroir of Cannabis: Terpene Metabolomics as a Tool to Understand Cannabis sativa Selections

The phytochemical diversity of Cannabis chemovars is not well understood, and many chemovars were created in informal breeding programs without records of parentage or the criteria for selection. Key criteria for selection sometimes included aroma notes and visual cues, which some breeders associated with pharmacological activity. We hypothesized that the process of selection for scents believed to be related to specific tetrahydrocannabinol levels has resulted in modified terpene biosynthesis in these chemovars. Thirty-two cannabinoids, 29 monoterpenes and 38 sesquiterpenes were measured in 33 chemovars from 5 licensed producers. A classification system based on cannabinoid content was used with targeted metabolomic tools to determine relationships in the phytochemistry. Three monoterpenes, limonene, β-myrcene, and α-pinene, and two sesquiterpenes, caryophyllene and humulene, were abundant in the majority of chemovars. Nine terpenes were present in tetrahydrocannabinol-dominant chemovars. Three monoterpenes and four sesquiterpenes were predominantly found in cannabidiol-containing chemovars. Low abundance terpenes may have been the aromatic cues identified by breeders. The medicinal activity of some of the terpenes is likely to contribute to the pharmacological effect of specific chemovars. Together, these data demonstrate the synergy of compounds in Cannabis chemovars and point to the need for additional research to understand the phytochemical complexity.

Pharmacological Foundations of Cannabis Chemovars.

An advanced Mendelian Cannabis breeding program has been developed utilizing chemical markers to maximize the yield of phytocannabinoids and terpenoids with the aim to improve therapeutic efficacy and safety. Cannabis is often divided into several categories based on cannabinoid content. Type I, Δ9-tetrahydrocannabinol-predominant, is the prevalent offering in both medical and recreational marketplaces. In recent years, the therapeutic benefits of cannabidiol have been better recognized, leading to the promotion of additional chemovars: Type II, Cannabis that contains both Δ9-tetrahydrocannabinol and cannabidiol, and cannabidiol-predominant Type III Cannabis. While high-Δ9-tetrahydrocannabinol and high-myrcene chemovars dominate markets, these may not be optimal for patients who require distinct chemical profiles to achieve symptomatic relief. Type II Cannabis chemovars that display cannabidiol- and terpenoid-rich profiles have the potential to improve both efficacy and minimize adverse events associated with Δ9-tetrahydrocannabinol exposure. Cannabis samples were analyzed for cannabinoid and terpenoid content, and analytical results are presented via PhytoFacts, a patent-pending method of graphically displaying phytocannabinoid and terpenoid content, as well as scent, taste, and subjective therapeutic effect data. Examples from the breeding program are highlighted and include Type I, II, and III Cannabis chemovars, those highly potent in terpenoids in general, or single components, for example, limonene, pinene, terpinolene, and linalool. Additionally, it is demonstrated how Type I - III chemovars have been developed with conserved terpenoid proportions. Specific chemovars may produce enhanced analgesia, anti-inflammatory, anticonvulsant, antidepressant, and anti-anxiety effects, while simultaneously reducing sequelae of Δ9-tetrahydrocannabinol such as panic, toxic psychosis, and short-term memory impairment.

Erratum: Evaluation of Cannabinoid and Terpenoid Content: Cannabis Flower Compared to Supercritical CO2 Concentrate.

A recent cannabis use survey revealed that 60% of cannabis users rely on smelling the flower to select their cannabis. Olfactory indicators in plants include volatile compounds, principally represented by the terpenoid fraction. Currently, medicinal- and adult-use cannabis is marketed in the United States with relatively little differentiation between products other than by a common name, association with a species type, and Δ-9 tetrahydrocannabinol/cannabidiol potency. Because of this practice, how terpenoid compositions may change during an extraction process is widely overlooked. Here we report on a comparative study of terpenoid and cannabinoid potencies of flower and supercritical fluid CO2 (SC-CO2) extract from six cannabis chemovars grown in Washington State. To enable this comparison, we employed a validated high-performance liquid chromatography/diode array detector methodology for quantification of seven cannabinoids and developed an internal gas chromatography-mass spectrometry method for quantification of 42 terpenes. The relative potencies of terpenoids and cannabinoids in flower versus concentrate were significantly different. Cannabinoid potency increased by factors of 3.2 for Δ-9 tetrahydrocannabinol and 4.0 for cannabidiol in concentrates compared to flower. Monoterpenes were lost in the extraction process; a ketone increased by 2.2; an ether by 2.7; monoterpene alcohols by 5.3, 7 and 9.4; and sesquiterpenes by 5.1, 4.2, 7.7, and 8.9. Our results demonstrate that the product of SC-CO2 extraction may have a significantly different chemotypic fingerprint from that of cannabis flower. These results highlight the need for more complete characterization of cannabis and associated products, beyond cannabinoid content, in order to further understand health-related consequences of inhaling or ingesting concentrated forms.

Cannabis Pharmacology: The Usual Suspects and a Few Promising Leads.

The golden age of cannabis pharmacology began in the 1960s as Raphael Mechoulam and his colleagues in Israel isolated and synthesized cannabidiol, tetrahydrocannabinol, and other phytocannabinoids. Initially, THC garnered most research interest with sporadic attention to cannabidiol, which has only rekindled in the last 15 years through a demonstration of its remarkably versatile pharmacology and synergy with THC. Gradually a cognizance of the potential of other phytocannabinoids has developed. Contemporaneous assessment of cannabis pharmacology must be even far more inclusive. Medical and recreational consumers alike have long believed in unique attributes of certain cannabis chemovars despite their similarity in cannabinoid profiles. This has focused additional research on the pharmacological contributions of mono- and sesquiterpenoids to the effects of cannabis flower preparations. Investigation reveals these aromatic compounds to contribute modulatory and therapeutic roles in the cannabis entourage far beyond expectations considering their modest concentrations in the plant. Synergistic relationships of the terpenoids to cannabinoids will be highlighted and include many complementary roles to boost therapeutic efficacy in treatment of pain, psychiatric disorders, cancer, and numerous other areas. Additional parts of the cannabis plant provide a wide and distinct variety of other compounds of pharmacological interest, including the triterpenoid friedelin from the roots, canniprene from the fan leaves, cannabisin from seed coats, and cannflavin A from seed sprouts. This chapter will explore the unique attributes of these agents and demonstrate how cannabis may yet fulfil its potential as Mechoulam's professed "pharmacological treasure trove."