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Cannabinoids are involved in physiological and neuromodulatory processes through their interactions with the human cannabinoid receptor-based endocannabinoid system. Their association with neurodegenerative diseases and brain reward pathways underscores the importance of evaluating and modulating cannabinoid activity for both understanding physiological mechanisms and developing therapeutic drugs. The use of agonists and antagonists could be strategic approaches for modulation. In this study, we introduce a bioelectronic sensor designed to monitor cannabinoid binding to receptors and assess their agonistic and antagonistic properties. We produced human cannabinoid receptor 1 (hCB1R) via an Escherichia coli expression system and incorporated it into nanodiscs (NDs). These hCB1R-NDs were then immobilized on a single-walled carbon nanotube field-effect transistor (swCNT-FET) to construct a bioelectronic sensing platform. This novel system can sensitively detect the cannabinoid ligand anandamide (AEA) at concentrations as low as 1 fM, demonstrating high selectivity and real-time response. It also successfully identified the hCB1R agonist Δ 9 -tetrahydrocannabinol and observed that the hCB1R antagonist rimonabant diminished the sensor signal upon AEA binding, indicating the antagonism-based modulation of ligand interaction. Consequently, our bioelectronic sensing platform holds potential for ligand detection and analysis of agonism and antagonism., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Glucose-6-phosphate dehydrogenase (G6PD) is a pentose phosphate pathway (PPP) enzyme that generates NADPH, which is required for cellular redox equilibrium and reductive biosynthesis. It has been demonstrated that abnormal G6PD activation promotes cancer cell proliferation and metastasis. To date, no G6PD inhibitor has passed clinical testing successfully enough to be launched as a medicine. As a result, in this investigation, cannabinoids were chosen to evaluate their anticancer potential by targeting G6PD. Molecular docking indicated that three molecules, Tetrahydrocannabinolic acid (THCA), Cannabichromenic acid (CBCA), and tetrahydrocannabivarin (THCV), have the highest binding affinities for G6PD of -8.61, -8.39, and 8.01 Kcal mol. ADMET analysis found that all of them were safe prospective drug candidates. Molecular dynamics (MD) simulation and MM-PBSA analysis confirm the structural compactness and lower conformational variation of protein-ligand complexes, thereby maintaining structural stability and rigidity. Thus, our in silico investigation exhibited all three cannabinoids as potential competitive inhibitors of G6PD., (© 2024 Wiley-VHCA AG, Zurich, Switzerland.)

Glycine receptors (GlyR) are regulated by small-molecule binding at several allosteric sites. Cannabinoids like tetrahydrocannabinol (THC) and N-arachidonyl-ethanol-amide (AEA) potentiate the GlyR response but their mechanism of action is not fully established. By combining millisecond coarse-grained (CG) MD simulations powered by Martini 3 with backmapping to all-atom representations, we have characterized the cannabinoid-binding site(s) at the zebrafish GlyR-α1 active state with atomic resolution. Based on hundreds of thousand ligand-binding events, we find that cannabinoids bind to the transmembrane domain of the receptor at both intrasubunit and intersubunit sites. For THC, the intrasubunit binding mode predicted in simulation is in excellent agreement with recent cryo-EM structures, while intersubunit binding recapitulates in full previous mutagenesis experiments. Intriguingly, AEA is predicted to bind at the same intersubunit site despite the strikingly different chemistry. Statistical analyses of the ligand-receptor interactions highlight potentially relevant residues for GlyR potentiation, offering experimentally testable predictions. The predictions for AEA have been validated by electrophysiology recordings of rationally designed mutants. The results highlight the existence of multiple cannabinoid-binding sites for the allosteric regulation of GlyR and put forward an effective strategy for the identification and structural characterization of allosteric binding sites., (© 2024. The Author(s).)

Cannabis sativa is known for producing over 120 distinct phytocannabinoids, with Δ 9 -tetrahydrocannabinol (Δ 9 -THC) and cannabidiol (CBD) being the most prominent, primarily in their acidic forms. Beyond Δ 9 -THC and CBD, a wide array of lesser-known phytocannabinoids, along with terpenes, flavonoids, and alkaloids, demonstrate diverse pharmacological activities, interacting with the endocannabinoid system (eCB) and other biological pathways. These compounds, characterized by phenolic structures and hydroxyl groups, possess lipophilic properties, allowing them to cross the blood-brain barrier (BBB) effectively. Notably, their antioxidant, anti-inflammatory, and neuro-modulatory effects position them as promising agents in treating neurodegenerative disorders. While research has extensively examined the neuropsychiatric and neuroprotective effects of Δ 9 -THC, other minor phytocannabinoids remain underexplored. Due to the well-established neuroprotective potential of CBD, there is growing interest in the therapeutic benefits of non-psychotropic minor phytocannabinoids (NMPs) in brain disorders. This review highlights the emerging research on these lesser-known compounds and their neuroprotective potential. It offers insights into their therapeutic applications across various major neurological conditions.

A multi-analyte liquid chromatography-tandem mass spectrometry (LC-MS/MS) method is described, involving the separation of delta-9-tetrahydrocannabinol (delta-9-THC) and delta-8-THC in addition to other commonly encountered drugs and metabolites. Briefly, sample preparation involved an alkaline liquid-liquid extraction (methyl tert-butyl ether) of blood (100 μl). The solvent layer was transferred, evaporated to dryness, reconstituted, and samples then separated on an Agilent Poroshell 120 EC-C18 100 Å (50 mm × 3.0 mm, 2.7 μm) analytical column using a multi-step gradient elution of 50 mM ammonium formate in water (pH 3.5) and 0.1% formic acid in methanol over 14 min. A SCIEX Triple Quad 6500+ system operating in scheduled multiple reaction monitoring and positive electrospray ionization was used for detection. There were no interferences, and matrix effects were generally acceptable (±20% of neat response). Linearity was achieved within the calibration range, including methylamphetamine (MA) (10-1000 ng/ml), 3,4-methylenedioxy-N-methylamphetamine (MDMA) (10-1,000 ng/ml), cocaine (10-1000 ng/ml), and two THC isomers (1-100 ng/ml). Accuracies of MA, MDMA, cocaine, and two THC isomers were 3.6 to 8.9%, -1.2 to 4%, -5.3 to 5.8%, and -11 to 14%, respectively; while precision estimates of the same were 1.6 to 5.4%, 1.7 to 5.3%, 1.2 to 4.5%, and 2 to 10%, respectively. Autosampler stability and dilution integrity were within acceptable limits, and no carryover was detected at the limit of detection. This validated LC-MS/MS method made the routine identification of both delta-9-THC and delta-8-THC in blood possible., (© 2023 The Authors. Drug Testing and Analysis published by John Wiley & Sons Ltd.)

Background: This article highlights the formulation of a solid Δ 9 -tetrahydrocannabinol (THC)-loaded ingestible prepared from pure THC distillate. Methods: A THC-containing ethanol-assisted cannabinoid nanoemulsion (EACNE) was created using a solvent displacement technique. Subsequently, the EACNE was converted to a solid powdery material while still retaining its THC potency, a format uniquely suited for "microdosing" applications. Results: EACNE had an average lipid droplet size of ∼190 nm, with a polydispersity index of 0.15, and an average droplet ζ potential of -49±10 mV. The nanoemulsion (NE) was colloidally stable for at least 6 weeks, with no meaningful change in cannabinoid potency over the experimental period, as determined by high-performance liquid chromatography analysis. The EACNE remained stable when subjected to physical stresses such as heat, freeze/thaw cycles, carbonation, dilution to beverage concentrations, high sucrose concentrations, and a pH range between 5 and 8. The microencapsulated EACNE demonstrated limited free-flowing behavior but was freely redispersible in water without any visible phase separation. Conclusions: We report the design, creation, and characterization of a THC NE generated without the use of specialized equipment, such as a microfluidizer or a high-pressure homogenizer. This emulsion could readily be converted to a water-redispersible powder. This embodiment is particularly suited for THC "microdosing," a practice that might decouple the health benefits of THC from its psychotropic effects.

Besides many other uses, dried Cannabis may be used for "tea" preparation. This study focused on a comprehensive characterization of an aqueous infusion prepared according to a common practice from three fairly different Cannabis cultivars. The transfer of 42 phytocannabinoids and 12 major bioactive compounds (flavonoids) into the infusion was investigated using UHPLC-HRMS/MS. Phytocannabinoid acids were transferred generally in a higher extent compared to their counterparts; in the case of Δ 9 -THC, it was only in the range of 0.4-1.9% of content in the Cannabis used. A dramatic increase of phytocannabinoids, mainly of the neutral species, occurred when cream was added during steeping, and the transfer of Δ 9 -THC into "tea" achieved a range of 53-64%. Under such conditions, drinking a 250 mL cup of such tea by a 70 kg person might lead to multiple exceedance of the Acute Reference Dose (ARfD), 1 μg/kg b.w., even in the case when using hemp with a Δ 9 -THC content below 1% in dry weight for preparation.

The adsorption of 11-nor-9-carboxy-∆ 9 -tetrahydrocannabinol (THC-COOH) by the suspended particles in sewage makes it fail to accurately monitor cannabis abuse. In this work, the model sewage sample was prepared through equivalent mixing the sewage from 10 different sewage treatment plants in Guangdong province of China and used as a comprehensive representative for investigating the adsorption and release behavior of THC-COOH on the suspended particles under different temperature and pH. The solid-liquid distribution of THC-COOH in sewage depended strongly on the adsorption and release properties which were susceptible to the temperature and pH, specially adjusting pH to 11.0 could release more than 90 % of THC-COOH from the suspended particles. By means of the kinetics models, pseudo-second-order kinetic and Weber-Morris models revealed the mechanism of adsorption and release of THC-COOH in sewage that was a relatively reversible and controllable process with multiple interactions, and then it was further confirmed by the validation experiment in a variety of actual sewage samples. According to the suggested pH, the modification of the existing detection protocol prior to high performance liquid chromatography-tandem triple quadrupole mass spectrometry (HPLC-TQ-MS/MS), was successfully applied to determination of THC-COOH in the stimulated positive samples, and the recoveries and RSDs were respectively 95.48-99.79 % and 4.0-5.6 %. The finding could greatly help improving the accuracy of not only the detection of THC-COOH in sewage but also the estimation data of the consumption level of cannabis in the related regions., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Gas chromatography-mass spectrometry (GC-MS) is widely used for the identification of cannabinoids in seized plant material. Conditions used for instrumental analysis should maximize decarboxylation, while minimizing the in situ production of Δ9-THC inside the GC inlet. In this study, decarboxylation of the acidic Δ9-THC precursor and in situ degradation of cannabidiol (CBD) were investigated using seven commercial GC liners with different deactivation chemistries and geometries. While the inlet temperature was previously optimized at 250°C in a previously validated assay, we systematically examined the temperature-dependent decarboxylation of tetrahydrocannabinolic acid-A (Δ9-THCA-A) and cyclization of CBD between 230°C and 310°C using different liners using favorable and unfavorable conditions. Significant differences in decarboxylation rate and CBD cyclization were observed between different liner types. While no temperature-dependent differences in decarboxylation rate were observed within liner type, liner-dependent differences were observed (α = 0.05), particularly between those with different geometry. In contrast, temperature and liner-dependent differences were observed for in situ formation of Δ9-THC (α = 0.05). This was influenced by liner geometry and to a smaller extent by surface deactivation. Effects were exacerbated with liner usage. While significant differences were observed using new and used GC liners, differences between liners of the same type but different lot numbers were not observed. Inter-instrument differences using the same liner were also evaluated and had minimal effect. Liner- and temperature-dependent effects were also confirmed using more than 20 cannabis plant extracts. Careful selection of liner, inlet conditions, and regular preventive maintenance can mitigate the risks associated with in situ formation Δ9-THC from CBD., (© 2023 John Wiley & Sons, Ltd.)

In this work, we investigated the ability of an electrochemical sensor to recognize Cannabis sativa L. samples with different total content of Δ 9 -tetrahydrocannabinol (Δ 9 -THC), determined by the levels of the psychoactive cannabinoid and of its biosynthetic precursor Δ 9 -tetrahydrocannabinolic acid (Δ 9 -THCA), using a multivariate approach. The voltammetric responses recorded with screen-printed electrodes modified with carbon black reflected the compositional differences from the different samples, in terms of cannabinoids of the vegetal material. PLS-DA models allowed for the correct classification of most C. sativa samples into the classes of legal and illegal samples according to total Δ 9 -THC content, based on threshold limits defined by the EU/US (0.3 % w/w) and Italian (0.6 % w/w) regulations. Satisfactory results were achieved in both cases, obtaining classification efficiency values in prediction of the external test set equal to 85 % and 100 % for the EU/US and Italian thresholds, respectively. The obtained results suggest the possibility to consider the proposed method as a starting point for the implementation of an automated device for rapid prescreening of total Δ 9 -THC content directly on site., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Cannabis sativa L. is a plant belonging to the Cannabaceae family known primarily for its recreational use due to the psychoactive properties of Δ 9 -tetrahydrocannabinol (THC). Despite this, several compounds belonging to the category of phytocannabinoids have shown in recent years a number of potentially promising therapeutic effects that have increased the interest in the pharmaceutical field towards this plant. However, the content of these compounds is very variable and influenced by different factors, such as growing conditions and time of the year. An indication of the status and age of Cannabis samples is provided by the content of CBN, a minor phytocannabinoid and degradation product of other phytocannabinoids, including THC. In this research work an innovative, solid state analytical approach has been developed to observe and evaluate the variations in the content of two phytocannabinoids (CBN and CBD) in Cannabis-derived products over time. In order to simulate the ageing of the Cannabis samples, an artificially accelerated ageing procedure has been developed and optimised by using high temperatures. The analyses were carried out using an innovative ATR-FTIR method for solid state analysis, enabling direct analysis of a solid sample without any pretreatment phase. This study has allowed the development of an innovative analytical approach for the evaluation of the age and state of conservation of Cannabis samples and may be a useful tool both in the industrial, pharmaceutical and forensic fields., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Cannabidiol (CBD) and Δ9-tetrahydrocannabinol (THC), the main components of Cannabis sativa plants, have attracted a significant amount of attention due to their biological activities. This study identified GPR18 as the target of partial agonist CBD activating the p42/p44 MAPK pathway leading to migration of endometrial epithelial cells. Induced fit docking (IFD) showed that the affinity of THC for GPR18 is higher than that of CBD, and molecular dynamics (MD) simulations showed that CBD-GPR18 complexes at 130/200 ns might have stable conformations, potentially activating GPR18 by changing the distances of key residues in its active pocket. In contrast, THC maintains "metastable" conformations, generating a "shrinking space" leading to full agonism of THC by adding mechanical constraints in GPR18's active pocket. Steered molecular dynamics (SMD) revealed GPR18's active pocket was influenced more by CBD's partial agonism compared with THC. This combined IFD-MD-SMD method may be used to explain the mechanism of activation of partial or full agonists of GPR18.

Cannabis is cultivated for therapeutic and recreational purposes where delta-9 tetrahydrocannabinol (THC) is a main target for its therapeutic effects. As the global cannabis industry and research into cannabinoids expands, more efficient and cost-effective analysis methods for determining cannabinoid concentrations will be beneficial to increase efficiencies and maximize productivity. The utilization of machine learning tools to develop near-infrared (NIR) spectroscopy-based prediction models, which have been validated from accurate and sensitive chemical analysis, such as gas chromatography (GC) or liquid chromatography mass spectroscopy (LCMS), is essential. Previous research on cannabinoid prediction models targeted decarboxylated cannabinoids, such as THC, rather than the naturally occurring precursor, tetrahydrocannabinolic acid (THCA), and utilize finely ground cannabis inflorescence. The current study focuses on building prediction models for THCA concentrations in whole cannabis inflorescences prior to harvest, by employing non-destructive screening techniques so cultivators may rapidly characterize high-performing cultivars for chemotype in real time, thus facilitating targeted optimization of crossbreeding efforts. Using NIR spectroscopy and LCMS to create prediction models we can differentiate between high-THCA and even ratio classes with 100% prediction accuracy. We have also developed prediction models for THCA concentration with a R 2 = 0.78 with a prediction error average of 13%. This study demonstrates the viability of a portable handheld NIR device to predict THCA concentrations on whole cannabis samples before harvest, allowing the evaluation of cannabinoid profiles to be made earlier, therefore increasing high-throughput and rapid capabilities.

Cultivation of industrial low-Δ 9 -tetrahydrocannabinol (Δ 9 -THC) hemp has created an oversupply of cannabidiol (CBD)-rich products. The fact that phytocannabinoids, including CBD, can be used as precursors to synthetically produce a range of THC variants-potentially located in a legal loophole-has led to a diversification of cannabis recreational drug markets. 'Hemp-compliant', 'hemp-derived' and 'semisynthetic' cannabinoid products are emerging and being advertised as (legal) alternatives for Δ 9 -THC. This study included a large panel (n = 30) of THC isomers, homologs, and analogs that might be derived via semisynthetic procedures. As a proxy for the abuse potential of these compounds, we assessed their potential to activate the CB 1 cannabinoid receptor with a β-arrestin2 recruitment bioassay (picomolar-micromolar concentrations). Multiple THC homologs (tetrahydrocannabihexol, THCH; tetrahydrocannabiphorol, THCP; tetrahydrocannabinol-C8, THC-C8) and THC analogs (hexahydrocannabinol, HHC; hexahydrocannabiphorol, HHCP) were identified that showed higher potential for CB 1 activation than Δ 9 -THC, based on either higher efficacy (E max ) or higher potency (EC 50 ). Structure-activity relationships were assessed for Δ 9 -THC and Δ 8 -THC homologs encompassing elongated alkyl chains. Additionally, stereoisomer-specific differences in CB 1 activity were established for various THC isomers (Δ 7 -THC, Δ 10 -THC) and analogs (HHC, HHCP). Evaluation of the relative abundance of 9(S)-HHC and 9(R)-HHC epimers in seized drug material revealed varying epimeric compositions between batches. Increased abundance of the less active 9(S)-HHC epimer empirically resulted in decreased potency, but sustained efficacy for the resulting diastereomeric mixture. In conclusion, monitoring of semisynthetic cannabinoids is encouraged as the dosing and the relative composition of stereoisomers can impact the harm potential of these drugs, relative to Δ 9 -THC products., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Phytocannabinoids with seven-carbon alkyl chains (phorols) have gained a lot of attention, as they are commonly believed to be more potent versions of typical cannabinoids with shorter alkyl chains. At the time of this article, cannabidiphorol (CBDP) and tetrahydrocannabiphorol (THCP) can both be purchased in the North American market, even though their biological activities are nearly unknown. To investigate their relative potency, we conducted in vitro receptor-binding experiments with CBDP (cannabinoid CB1/CB2 receptor antagonism, serotonin 5HT-1A agonism, dopamine D2S (short form) agonism, and mu-opioid negative allosteric modulation) and compared the observed activity with that of CBD. To our knowledge, this is the first publication to investigate CBDP's receptor activity in vitro. A similar activity profile was observed for both CBD and CBDP, with the only notable difference at the CB2 receptor. Contrary to common expectations, CBD was found to be a slightly more potent CB2 antagonist than CBDP ( p < 0.05). At the highest tested concentration, CBD demonstrated antagonist activity with a 33% maximum response of SR144528 (selective CB2 antagonist/inverse agonist). CBDP at the same concentration produced a weaker antagonist activity. A radioligand binding assay revealed that among cannabinoid and serotonin receptors, CB2 is likely the main biological target of CBDP. However, both CBD and CBDP were found to be significantly less potent than SR144528. The interaction of CBDP with the mu-opioid receptor (MOR) produced unexpected results. Although the cannabidiol family is considered to be a set of negative allosteric modulators (NAMs) of opioid receptors, we observed a significant increase in met-enkephalin-induced mu-opioid internalization when cells were incubated with 3 µM of CBDP and 1 µM met-enkephalin, a type of activity expected from positive allosteric modulators (PAMs). To provide a structural explanation for the observed PAM effect, we conducted molecular docking simulations. These simulations revealed the co-binding potential of CBDP (or CBD) and met-enkephalin to the MOR.

The 2018 Farm Bill defines marijuana as Cannabis sativa L. or any derivative thereof that contains greater than 0.3% Δ 9 -tetrahydrocannabinol (Δ 9 -THC) on a dry weight basis. The main cannabinoids present in Cannabis sativa L., Δ 9 -THC and cannabidiol (CBD), are structural isomers that cannot be differentiated using direct mass spectrometry with soft ionization techniques alone. Due to the classification of marijuana as a Schedule I controlled substance, the differentiation of Δ 9 -THC and CBD is crucial within the seized drug community. This study explores the use of Ag-ligand ion complexation and electrospray ionization tandem mass spectrometry (ESI-MS/MS) for the differentiation of Δ 9 -THC and CBD using six different Ag complexes. Differences between the binding affinities of Δ 9 -THC and CBD for [Ag(PPh 3 )(OTf)] 2 lead to the formation of unique product ions at m / z 421/423, m / z 353/355, and m / z 231 for CBD, enabling the differentiation of CBD from Δ 9 -THC. When applied to the analysis of known Δ 9 -THC:CBD mixture ratios, the developed [Ag(PPh 3 )(OTf)] 2 ion complexation method was able to differentiate Δ 9 -THC-rich and CBD-rich samples based on the average abundance of the product ions at m / z 421/423. The developed approach was then applied to methanolic extracts of 20 authentic cannabis samples with known Δ 9 -THC and CBD compositions, resulting in a 95% correct classification rate. Even though the developed Ag-ligand ion complexation method was only demonstrated for the qualitative differentiation of Δ 9 -THC-rich and CBD-rich cannabis, this study establishes a foundation for the use of Ag-ligand ion complexation that is essential for future quantitative approaches.

Δ 9 -THC, the psychotropic cannabinoid in Cannabis sativa L., for many years has been the focus of all the pharmacological attention as the main promising principle of the plant. Recently, however, cannabidiol (CBD) has brought a sudden change in the scenario, exponentially increasing the interest in pharmacology as the main non-psychotropic cannabinoid with potential therapeutic, cosmetical and clinical applications. Although the reactivity of CBD and Δ 9 -THC has been considered, little attention has been paid to the possible photodegradation of these cannabinoids in the vegetal matrix and the data available in the literature are, in some cases, contradictory. The aim of the present work is to provide a characterization of the photochemical behaviour of CBD and Δ 9 -THC in three cannabis chemotypes, namely I (Δ 9 -THC 2.50% w/w ), II (CBD:Δ 9 -THC 5.82% w/w :3.19% w/w ) and III (CBD 3.02% w/w )., (© 2024. The Author(s).)

The human orphan G protein-coupled receptor GPR18, activated by Δ 9 -tetrahydrocannabinol (THC), constitutes a promising drug target in immunology and cancer. However, studies on GPR18 are hampered by the lack of suitable tool compounds. In the present study, potent and selective GPR18 agonists were developed showing low nanomolar potency at human and mouse GPR18, determined in β-arrestin recruitment assays. Structure-activity relationships were analyzed, and selectivity versus cannabinoid (CB) and CB-like receptors was assessed. Compound 51 (PSB-KK1415, EC 50 19.1 nM) was the most potent GPR18 agonist showing at least 25-fold selectivity versus CB receptors. The most selective GPR18 agonist 50 (PSB-KK1445, EC 50 45.4 nM) displayed >200-fold selectivity versus both CB receptor subtypes, GPR55, and GPR183. The new GPR18 agonists showed minimal species differences, while THC acted as a weak partial agonist at the mouse receptor. The newly discovered compounds represent the most potent and selective GPR18 agonists reported to date.

Cannabis sativa is known for its therapeutic benefit in various diseases including pain relief by targeting cannabinoid receptors. The primary component of cannabis, Δ9-tetrahydrocannabinol (THC), and other agonists engage the orthosteric site of CB1, activating both Gi and β-arrestin signaling pathways. The activation of diverse pathways could result in on-target side effects and cannabis addiction, which may hinder therapeutic potential. A significant challenge in pharmacology is the design of a ligand that can modulate specific signaling of CB1. By leveraging insights from the structure-function selectivity relationship (SFSR), we have identified Gi signaling-biased agonist-allosteric modulators (ago-BAMs). Further, two cryoelectron microscopy (cryo-EM) structures reveal the binding mode of ago-BAM at the extrahelical allosteric site of CB1. Combining mutagenesis and pharmacological studies, we elucidated the detailed mechanism of ago-BAM-mediated biased signaling. Notably, ago-BAM CB-05 demonstrated analgesic efficacy with fewer side effects, minimal drug toxicity and no cannabis addiction in mouse pain models. In summary, our finding not only suggests that ago-BAMs of CB1 provide a potential nonopioid strategy for pain management but also sheds light on BAM identification for GPCRs., Competing Interests: Competing interests statement:The authors declare no competing interest.

This work developed an aptamer-dye complex as a label-free ratiometric fluorescence sensor for rapid analysis of THC and its metabolite in sewage samples. Integrated with a portable fluorescence capture device, this sensor exhibited excellent sensitivity with visualization of as low as 0.6 μM THC via naked-eye observation, and THC analysis can be accomplished within 4 min, which would be a complementary tool for quantifying THC in sewage samples to estimate cannabis consumption.

Since around 2021, products claiming to contain a Δ 9 -tetrahydrocannabinol (THC) analog with different lengths of alkyl chain at C-3 position have been sold on the internet in Japan. Δ 9 -THC has a pentyl group derived from the precursor olivetol at the C-3 position. These products include liquid cartridges for electronic cigarettes, herbal products, and gummy products. This study analyzed and determined the ingredients in five oil products distributed on the internet from 2022 to 2023 that claim to contain THC analogs. Samples of each product were used for GC-MS and LC-MS measurements. After isolating and purifying the unknown components from the products, structural analysis was performed by measuring 1 H, 13 C-NMR and various two-dimensional NMR [HH correlation spectroscopy (H-H COSY), heteronuclear multiple quantum coherence (HMQC), heteronuclear multiple-bond correlation (HMBC), and nuclear Overhauser effect spectroscopy (NOESY)]. The analysis identified Δ 8 -tetrahydrocannabivarin (THCV), Δ 9 -THCV, Δ 8 -tetrahydrocannabutol (THCB), Δ 9 -THCB, Δ 8 -tetrahydrocannabihexol (THCH), Δ 9 -THCH, Δ 8 -3-octyl-THC (THCjd) and Δ 9 -THCjd. These compounds were Δ 8 -THC or Δ 9 -THC analogs with different lengths of alkyl chain at C-3 position. Meanwhile, Δ 4(8) -iso-THCV and Δ 11 -THCB were identified as minor components of the product, and were considered to be the reaction byproducts of the synthesis of the Δ 8 -THC or Δ 9 -THC analogs. In the future, there are concerns about the distribution of products containing new THC analogs. Therefore, continuous provision monitoring of newly detected in the products is important.

Sativex is a cannabis-based medicine that comes in the form of an oromucosal spray. It contains equal amounts of Δ9-tetrahydrocannabinol and cannabidiol, two compounds derived from cannabis plants. Sativex has been shown to have positive effects on symptoms of amyotrophic lateral sclerosis (ALS), multiple sclerosis (MS), and sleep disorders. It also has analgesic, antiinflammatory, antitumoral, and neuroprotective properties, which make it a potential treatment option for other neurological disorders. The article reviews the results of recent preclinical and clinical studies that support the therapeutic potential of Sativex and the molecular mechanisms behind its neuroprotective benefits in various neurological disorders. The article also discusses the possible advantages and disadvantages of using Sativex as a neurotherapeutic agent, such as its safety, efficacy, availability, and legal status., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Genetic markers can represent a valuable tool for forensic purposes in discriminating between fiber-type and drug-type cannabis. The aim of this research was to evaluate developed genetic markers for tetrahydrocannabinolic acid synthase (THCAS) when applied on certified hemp (14 varieties) and forensic casework samples of four chemotypes (40 seizures). Chemotype-associated PCR-based markers did not enable reliable selective amplification despite the difference in cannabinoid composition. In order to characterize forensic samples of unknown origin, THCAS sequencing was performed. The comparison of THCAS sequences, including additional accessions, indicated high genetic similarity of hemp varieties. Confiscated samples of intermediate, THC, CBD and CBG type were clearly separated from fiber-type accessions and assigned to drug-type cluster. Despite the unknown origin, their position on the tree support the notion that they are more related to drug-type accessions than to the fiber-type. However, no clear distinction between chemotypes was found. Furthermore, 26 amino acid substitutions were revealed in THCAS that clearly separate hemp varieties and neither of them cluster with any other tested sample., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Tetrahydrocannabinol (THC) and cannabidiol (CBD) are the two "major cannabinoids". However, their incorporation into clinical and nutraceutical preparations is challenging, owing to their limited bioavailability, low water solubility, and variable pharmacokinetic profiles. Understanding the organic chemistry of the major cannabinoids provides us with potential avenues to overcome these issues through derivatization. The resulting labile pro-drugs offer ready cannabinoid release in vivo , have augmented bioavailability, or demonstrate interesting pharmacological properties in their own right. This review identifies and discusses a subset of these advanced derivatization strategies for the major cannabinoids, where the starting material is the pure phytocannabinoid itself, and the final product either a cannabinoid pro-drug, or a novel pharmacoactive material.

Gas chromatography (GC) techniques for analyzing and determining the cannabinoid profile in cannabis ( Cannabis sativa L.) are widely used in standard laboratories; however, these methods may mislabel the profile when used under rapid conditions. Our study aimed to highlight this problem and optimize GC column conditions and mass spectrometry (MS) parameters to accurately identify cannabinoids in both standards and forensic samples. The method was validated for linearity, selectivity, and precision. It was observed that when tetrahydrocannabinol (Δ9-THC) and cannabidiolic acid (CBD-A) were examined using rapid GC conditions, the resulting derivatives generated identical retention times. Wider chromatographic conditions were applied. The linear range for each compound ranged from 0.02 μg/mL to 37.50 μg/mL. The R 2 values ranged from 0.996 to 0.999. The LOQ values ranged from 0.33 μg/mL to 5.83 μg/mL, and the LOD values ranged from 0.11 μg/mL to 1.92 μg/mL. The precision values ranged from 0.20% to 8.10% RSD. In addition, forensic samples were analyzed using liquid chromatography (HPLC-DAD) in an interlaboratory comparison test, with higher CBD and THC content than GC-MS determination ( p < 0.05) in samples. Overall, this study highlights the importance of optimizing GC techniques to avoid mislabeling cannabinoids in cannabis samples.

In Cannabis sativa L. the presence of delta 9-tetrahydrocannabinolic acid (THCA) above legal limit is a challenging issue that still restricts the industrial exploitation of this promising crop. In recent years, the interest of entrepreneurs and growers who see hemp as a dynamic and profitable crop was joined by the growing knowledge on C. sativa genetics and genomics, accelerated by the application of high throughput tools. Despite the renewed interest in the species, much remains to be clarified, especially about the long-standing problem of THCA in hemp inflorescences, which could even result in the seizure of the whole harvest. Although several hypotheses have been formulated on the accumulation of this metabolite in industrial varieties, none is conclusive yet. In this work, individuals of a population of the hemp cultivar 'FINOLA' obtained from commercial seeds were investigated for total THC level and examined at molecular level. A marker linked to THCA synthase was found at a high incidence in both male and female plants, suggesting a considerable genetic variability within the seed batch. Full-length sequences encoding for putatively functional THCA synthases were isolated for the first time from the genome of both female and male plants of an industrial hemp variety and, using transcriptional analysis, the THCA synthase expression was quantified in mature inflorescences of individuals identified by the marker. Biochemical analyses finally demonstrated for these plants a 100% association between the predicted and actual chemotype., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

There appears to be consensus among Cannabis biologists that delta-9-tetrahydrocannabinolic acid A (THCA-A) is the exclusive product of the enzyme THCA synthase. This then leaves an open question for formation of the THCA-A structural isomer, delta-9-tetrahydrocannabinolic acid B (THCA-B), discovered as a minor product in Cannabis in 1969. With no reasonable biochemical pathway to explain the presence of THCA-B in Cannabis , a synthetic route was next considered. Using established literature precedent, a photochemical mechanism has been proposed for the conversion of THCA-A to cannabidiolic acid (CBDA), followed by conversion of CBDA to THCA-B employing bond-breaking and bond-forming reactions.

Although much is known about Δ9-tetrahydrocannabinol and its inactive open ring isomer, cannabidiol, far less is known about the effects, metabolism, and pharmacodynamics of Δ9-tetrahydrocannabinol's double-bond isomer, Δ8-tetrahydrocannabinol. With the passage of the so-called United States "Farm Bill," which was made law in order to allow legal hemp cultivation in the United States, more needs to be known about the effects of Δ8-tetrahydrocannabinol, a double-bond isomer of Δ9-tetrahydrocannabinol, and cannabidiol (CBD), which is an open-ring isomer of Δ8-tetrahydrocannabinol. It is the aim of the review to summarize current knowledge of Δ8-tetrahydrocannabinol and CBD, including the pharmacodynamics and pharmacokinetics of CBD. Also, plant genetics, the effect of cannabinoids on the current topic of viral entry into mammalian cells, and the current practice of vaping, dabbing, and dripping are covered., (Copyright © 2023 Forensic Science Review.)

Introduction: Medical uses of Cannabis sativa L. have gained interest in recent decades, which highlights the need for defining appropriate quality specifications for Cannabis-based products. However, the complexity of plant matrices and structural similarity between cannabinoids make analytical development a challenging task. Thus, the application of analytical quality by design (AQbD)-driven approaches can favour the development of fit-for-purpose methods., Objectives: To develop a high-performance liquid chromatography diode array detector (HPLC-DAD) method for simultaneous quantification of cannabidiol, Δ 9 -tetrahydrocannabinol, cannabidiolic acid, tetrahydrocannabinolic acid, and cannabinol in C. sativa by applying an AQbD-driven approach., Materials and Methods: Critical method attributes (CMA) were established following the analytical target profile. Critical method variables (CMV) were categorised based on risk assessment and literature review. Selected CMV regarding sample preparation and chromatographic conditions were optimised using response surface methodology (RSM). The working point was estimated by multiple response optimisation using Deringer's desirability function. The validity of the optimal conditions was confirmed experimentally. Method validation was performed according to ANVISA and ICH guidelines. Relative response factors (RRFs) were also determined., Results and Discussion: Baseline resolution of 12 major cannabinoids was achieved in a 35 min chromatographic analysis. All experimental responses obtained during confirmatory analyses were within the prediction intervals (PI 95% ). Method's selectivity, linearity (10-100 μg/mL), precision, bias, extraction recovery, and ruggedness were satisfactorily demonstrated., Conclusions: The application of an AQbD-driven approach allowed for a better understanding of the effects of the ensemble of CMV on the analyte's behaviour, enabling the definition of appropriate conditions to ensure consistent achievement of the intended method's performance., (© 2022 John Wiley & Sons Ltd.)

There have been conflicting reports on the impact of Cannabis sativa impact on reproductive function. Hence this study was aimed to ascertain the impact of tetrahydrocannabinol (THC) and cannabidiol (CBD) binding affinity on human androgen receptor (AR) via computational molecular dynamic simulation. The human AR coordinate in this study is derived from human AR in complex with the ligand metribolone (R18) (PBD ID: 1E3G) template using (MODELER version. 9.15). CBD (PubChem CID: 644019), and THC (PubChem CID: 16078) 2D structures were retrieved from PubChem and docked (Autodock-Vina inbuilt in PyMol into the active site of human AR using the coordinates of the co-crystalized ligand (R18). All atomic representations in this study were created using visual molecular dynamics (VMD) tools. The result revealed that neither CBD nor THC bear significant 2D similarity with R18. Despite the diversity within the chemical space, both CBD and THC poses bond flexibility required to bind avidly to AR with the docking scores comparable to R18. In fully bound state, the three compounds engage the AR pocket hydrophobic residues such as L701, L704, and L707, and aromatic residues such as F764. Polar contacts with T877 observed in R18 bound state is avoided in the THC and CBD bound states. Moreso, the results revealed that CBD has lesser binding energy compared to THC and R18 compound which serves as standard. This study hypothesized that CBD and THC binds complimentarily to the pocket AR, indicating a likely inhibition of reproductive function and prostate cancer progression., (© 2022 Wiley-VCH GmbH.)

Cannabinoid receptor type 2 (CB2R) is a member of the class A G protein-coupled receptor (GPCRs) family and a component of the endocannabinoid system that is modulated by the psychoactive chemical from Cannabis sativa, partial agonist Δ 9 -tetrahydrocannabinol (Δ 9 -THC). Selective activation of CB2R allows for the treatment of inflammatory and immune-related conditions without the psychotropic effects of CB1R. While CB2R-selective agonists are available, CB2R partial agonists are scarce. Hence, the pharmacological difference between CB2R full agonists and partial agonists remains to be deciphered, prompting the search for novel partial agonists. Here, using an induced-fit docking approach, we built a partial agonist Δ 9 -THC bound CB2R system from the inactive CB2R structure (PDB ID: 5ZTY) and performed microsecond molecular dynamics (MD) simulations. The simulations reveal an upward shift of the "toggle switch" W6.48 (258) and minor outward movement of the transmembrane helix 6 (TM6). Dynamic network model identifies a possible communication path between the ligand and the toggle switch" W6.48 (258) . Furthermore, to identify potential CB2R partial agonists, we conducted structure-based virtual screening of ZINC15 "Druglike" library containing 17,900742 compounds against 3 conformations derived from MD simulation of CB2R complexed with partial agonist Δ 9 -THC using Glide virtual screening protocol comprising various filters with increasing accuracy. Nine diverse compounds predicted to have high MM-GBSA binding energy scores and good ADMET properties (including high gastrointestinal absorption and low toxicity) are proposed as potential CB2R partial agonists., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

The popularity of vaping cannabis products has increased sharply in recent years. In 2019, a sudden onset of electronic cigarette/vaping-associated lung injury (EVALI) was reported, leading to thousands of cases of lung illness and dozens of deaths due to the vaping of tetrahydrocannabinol (THC)-containing e-liquids that were obtained on the black market. A potential cause of EVALI has been hypothesized due to the illicit use of vitamin E acetate (VEA) in cannabis vape cartridges. However, the chemistry that modifies VEA and THC oil, to potentially produce toxic byproducts, is not well understood under different scenarios of use. In this work, we quantified carbonyls, organic acids, cannabinoids, and terpenes in the vaping aerosol of pure VEA, purified THC oil, and an equal volume mixture of VEA and THC oil at various coil temperatures (100-300 °C). It was found under the conditions of our study that degradation of VEA and cannabinoids, including Δ 9 -THC and cannabigerol (CBG), occurred via radical oxidation and direct thermal decomposition pathways. Evidence of terpene degradation was also observed. The bond cleavage of aliphatic side chains in both VEA and cannabinoids formed a variety of smaller carbonyls. Oxidation at the ring positions of cannabinoids formed various functionalized products. We show that THC oil has a stronger tendency to aerosolize and degrade compared to VEA at a given temperature. The addition of VEA to the e-liquid nonlinearly suppressed the formation of vape aerosol compared to THC oil. At the same time, toxic carbonyls including formaldehyde, 4-methylpentanal, glyoxal, or diacetyl and its isomers were highly enhanced in VEA e-liquid when normalized to particle mass.

Cannabinoid receptor 1 (CB 1 ) is a therapeutically relevant drug target for controlling pain, obesity, and other central nervous system disorders. However, full agonists and antagonists of CB 1 have been reported to cause serious side effects in patients. Therefore, partial agonists have emerged as a viable alternative as they can mitigate overstimulation and side effects. One of the key bottlenecks in the design of partial agonists, however, is the lack of understanding of the molecular mechanism of partial agonism itself. In this study, we examine two mechanistic hypotheses for the origin of partial agonism in cannabinoid receptors and predict the mechanistic basis of partial agonism exhibited by Δ 9 -Tetrahydrocannabinol (THC) against CB 1 . In particular, we inspect whether partial agonism emerges from the ability of THC to bind in both agonist and antagonist-binding poses or from its ability to only partially activate the receptor. We used extensive molecular dynamics simulations and Markov state modeling to capture the THC binding in both antagonist and agonist-binding poses in the CB 1 receptor. Furthermore, we predict that binding of THC in the agonist-binding pose leads to rotation of toggle switch residues and causes partial outward movement of intracellular transmembrane helix 6 (TM6). Our simulations also suggest that the alkyl side chain of THC plays a crucial role in determining partial agonism by stabilizing the ligand in the agonist and antagonist-like poses within the pocket. Taken together, this study provides important insights into the mechanistic origin of the partial agonism of THC., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Introduction: Cannabis sativa L. ( C. sativa ) is used since ancient times to produce fabrics, baskets, and cords. Later, different ethnic groups used to burn the leaves and flowers of psychotropic cultivars with high Δ9-tetrahydrocannabinol (D9-THC) levels, during the religious or propitiatory rites to alter the state of consciousness. To date, it is not known whether also nonpsychotropic cultivars of C. sativa were used during these rites, and whether these varieties could have an effect on human behavior. This study aimed to evaluate the behavioral effects of an extract of nonpsychotropic C. sativa (NP-CS) in mice. Materials and Methods: An extract of a nonpsychotropic cultivar of C. sativa dissolved in medium-chain triglyceride oil was used and the different phytochemical components were evaluated. The relative composition in terms of phytocannabinoid content was assessed by reverse phase high-performance liquid chromatography coupled to UV detection (RP-HPLC-UV), and the volatile components were analyzed by gas chromatography-mass spectrometry (GC-MS). In addition, the behavioral effect of NP-CS was assessed on a wild-type mouse model. The animals were treated for 14 days (oral gavage) and motility, anxiety, and social effects were assessed. Results: RP-HPLC-UV analysis demonstrated that D9-THC was present in lower concentration with respect to other cannabinoids, like cannabidiol. Furthermore, the GC-MS analysis revealed the presence of several terpenoids. Concerning in vivo studies, chronic treatment with NP-CS did not alter body weight, motility, and anxiety and increased social interaction. Conclusions: This study highlighted the prosocial effects of NP-CS.

"E-Cigarette (e-cig) Vaping-Associated Acute Lung Injury" (EVALI) has been linked to vitamin-E-acetate (VEA) and Δ-9-tetrahydrocannabinol (THC), due to their presence in patients' e-cigs and biological samples. Lacking standardized methodologies for patients' data collection and comprehensive physicochemical/toxicological studies using real-world-vapor exposures, very little data are available, thus the underlying pathophysiological mechanism of EVALI is still unknown. This review aims to provide a comprehensive and critical appraisal of existing literature on clinical/epidemiological features and physicochemical-toxicological characterization of vaping emissions associated with EVALI. The literature review of 161 medical case reports revealed that the predominant demographic pattern was healthy white male, adolescent, or young adult, vaping illicit/informal THC-containing e-cigs. The main histopathologic pattern consisted of diffuse alveolar damage with bilateral ground-glass-opacities at chest radiograph/CT, and increased number of macrophages or neutrophils and foamy-macrophages in the bronchoalveolar lavage. The chemical analysis of THC/VEA e-cig vapors showed a chemical difference between THC/VEA and the single THC or VEA. The chemical characterization of vapors from counterfeit THC-based e-cigs or in-house-prepared e-liquids using either cannabidiol (CBD), VEA, or medium-chain triglycerides (MCT), identified many toxicants, such as carbonyls, volatile organic compounds, terpenes, silicon compounds, hydrocarbons, heavy metals, pesticides and various industrial/manufacturing/automotive-related chemicals. There is very scarce published toxicological data on emissions from THC/VEA e-liquids. However, CBD, MCT, and VEA emissions exert varying degrees of cytotoxicity, inflammation, and lung damage, depending on puffing topography and cell line. Major knowledge gaps were identified, including the need for more systematic-standardized epidemiological surveys, comprehensive physicochemical characterization of real-world e-cig emissions, and mechanistic studies linking emission properties to specific toxicological outcomes.

Cannabis e-cigarettes containing Δ 8 -tetrahydrocannabinol (Δ 8 -THC) produced synthetically from hemp-derived cannabidiol (CBD) have recently risen in popularity as a legal means of cannabis consumption, but questions surrounding purity and unlabeled additives have created doubts of their safety. Herein, NMR, GC-MS, and ICP-MS were used to analyze major components of 27 products from 10 brands, and it was determined none of these had accurate Δ 8 -THC labeling, 11 had unlabeled cutting agents, and all contained reaction side-products including olivetol, Δ 4(8) - iso -tetrahydrocannabinol, 9-ethoxyhexahydrocannabinol, Δ 9 -tetrahydrocannabinol (Δ 9 -THC), heavy metals, and a novel previously undescribed cannabinoid, iso -tetrahydrocannabifuran.

The therapeutic effects of Δ 9 -tetrahydrocannabinol (Δ 9 -THC) can be enhanced by modifications of the pentyl moiety at C-3. The engineering of Cannabis sativa olivetolic acid cyclase and tetraketide synthase with F24I and L190G substitutions, respectively, in the biosynthesis of Δ 9 -THC serves as a platform for the generation of resorcylic acids up to 6-undecylresorcylic acid. These results provide insights into the development of THC analogs with chemically distinct acyl moieties at C-3.

Cannabidiol (CBD), a major non-psychoactive cannabinoid, has a lot of attention due to its potential relaxing properties and led the trend in commercial CBD aroma/oral hemp seed oil from the Japanese market. In this study, a routine assay for evaluating CBD oil samples was performed using LC coupled with tandem mass spectrometry (LC-MS/MS) and was used to apply the convertible tetrahydrocannabinol (THC) in acetic acid conditions. Based on the electrospray positive ion mode, the detection of cannabidiolic acid (CBDA; m/z 359 > 219), cannabigerolic acid (CBGA; m/z 361 > 343), cannabigerol (CBG; m/z 317 > 193), CBD (m/z 315 > 193), THC (m/z 315 > 193) and cannabinol (CBN; m/z 311 > 223) was performed by satisfying separation with high density of C 18 column. Oil samples (50 mg) were diluted with isopropanol (5 mL), to which stable isotope internal standards were added by dilution with methanol/water (50/50), and accuracy rates ranged from 97.8 to 102.2%. This method was used to evaluate the CBD oil products (5 kinds) from the Japanese market. Our survey found obvious counterfeit (non-detectable CBD) CBD oil from Japanese market. Following that, we investigated the conversion of THC in CBD oil samples in simple conditions such as 10% acetic acid and 70 °C for 6 h and discovered that converts THC proportions are approximately 5% ((THC content/CBD content) × 100) and <1.0%. Thus, our developed LC-MS/MS assay could be applied to monitor the CBD concentration and convertible THC from CBD oil.

The FDA approved drug Dronabinol was identified in a previous study applying virtual screening using the haemozoin crystal as a target against malaria parasites. The active ingredient of dronabinol is synthetic tetrahydrocannabinol (THC), which is one of the major cannabinoids from Cannabis sativa. Traditional use of cannabis for malaria fever was reported in the world's oldest pharmacopoeia, dating to around 5000 years ago. In this research we report that THC inhibits β-haematin (synthetic haemozoin) and malaria parasite growth. Due the psychoactivity of THC, CBD, the other major naturally occurring cannabinoid that lacks the off-target psychoactive effects of THC, was also tested and inhibited β-haematin but showed only a mild antimalarial activity. To evaluate whether THC inhibit haemozoin formation, we performed a cellular haem fractionation assay that indicated that is not the likely mechanism of action. For the first time, the cannabinoid chemical structure is raised as a new chemical class to be further studied for malaria treatment, aiming to overcome the undesirable psychoactive effects of THC and optimize the antimalarial effects., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

The outbreak of e-cigarette or vaping product use-associated lung injury (EVALI) has been cause for concern to the medical community, particularly given that this novel illness has coincided with the COVID-19 pandemic, another cause of severe pulmonary illness. Though cannabis e-cigarettes tainted with vitamin E acetate were primarily associated with EVALI, acute lung injuries stemming from cannabis inhalation were reported in the literature prior to 2019, and it has been suggested that cannabis components or additives other than vitamin E acetate may be responsible. Despite these concerning issues, novel cannabis vaporizer ingredients continue to arise, such as Δ 8 -tetrahydrocannabinol, Δ 10 -tetrahydrocannabinol, hexahydrocannabinol, and cannabichromene. In order to address cannabis e-cigarette safety and vaping in an effective manner, we provide a comprehensive knowledge of the latest products, delivery modes, and ingredients. This perspective highlights the types of cannabis vaping modalities common to the United States cannabis market, with special attention to cartridge-type cannabis e-cigarette toxicology and their involvement in the EVALI outbreak, in particular, acute lung injurious responses. Novel ingredient chemistry, origins, and legal statuses are reviewed, as well as the toxicology of known cannabis e-cigarette aerosol components.

Cannabis access laws allow for the use of cannabis in private and the trade, purchase and use of hemp-related products as a complementary medicine and for other benefits. Cannabidiol (CBD) has the treatment potential for several conditions but, with the lack of resources in South Africa to maintain the legislation, products contaminated with delta-9-tetrahydrocannabinol (Δ9-THC) are sold by some suppliers who do not comply with the legislative provisions in terms of the threshold concentrations for Δ9-THC. This dilemma complicates a medical review officer's decision regarding intentional use of Δ9-THC or otherwise, since a CBD user may have purchased the product legally and in good faith. Hemp- and CBD-containing products were analysed by gas chromatography-mass spectrometry and compliance was assessed for CBD and Δ9-THC purity against the legislative thresholds. A strategy based on metabolite ratios is suggested to distinguish between intentional or irresponsible cannabis use and legitimate CBD use.

The cis -stereoisomers of Δ 9 -THC [(-)- 3 and (+)- 3 ] were identified and quantified in a series of low-THC-containing varieties of Cannabis sativa registered in Europe as fiber hemp and in research accessions of cannabis. While Δ 9 - cis -THC ( 3 ) occurs in cannabis fiber hemp in the concentration range of (-)-Δ 9 - trans -THC [(-)- 1 ], it was undetectable in a sample of high-THC-containing medicinal cannabis. Natural Δ 9 - cis -THC ( 3 ) is scalemic (ca. 80-90% enantiomeric purity), and the absolute configuration of the major enantiomer was established as 6a S ,10a R [(-)- 3 ] by chiral chromatographic comparison with a sample available by asymmetric synthesis. The major enantiomer, (-)-Δ 9 - cis -THC [(-)- 3 ], was characterized as a partial cannabinoid agonist in vitro and elicited a full tetrad response in mice at 50 mg/kg doses. The current legal discrimination between narcotic and non-narcotic cannabis varieties centers on the contents of "Δ 9 -THC and isomers" and needs therefore revision, or at least a more specific wording, to account for the presence of Δ 9 - cis -THCs [(+)- 3 and (-)- 3 ] in cannabis fiber hemp varieties.

The present study shows the variability in chemical profiles of three different varieties of Cannabis sativa extracts used in medicinal cannabis oil when using two different extraction and evaporation methods. Procedures were compared by analyzing rate of total terpenoids to the principal cannabinoids, tetrahydrocannabinol (THC) and cannabidiol (CBD), using gas chromatography coupled to mass spectrometry (GC-MS). The extraction of inflorescences using soxhlet method showed the highest mass of final extract. Also, the use of heating and the presence of oxygen when evaporating the solvents, greatly modified the final profiles due to evaporation or chemical reactions. These variations in chemical profiles must be carefully taken into account and standardized in the elaboration of medicinal oils.

As the frequency of cannabis-based therapy increases, the ability to distinguish intake of cannabis-based medicines from recreational cannabis use becomes desirable. Minor cannabinoids have been suggested to indicate recreational cannabis use in biological matrices but are unreliable when presumably also present in directly plantderived medicines. Thus, for therapeutics such as medical cannabis, Sativex® and Dronabinol, a more thorough investigation of cannabinoid profiles is required to identify possible distinguishing markers. In this study, 16 phytocannabinoids were quantified in samples of seized and medical cannabis, Sativex® and Dronabinol from two different manufacturers, using a validated LC-MS/MS method. Analytes included delta-9- tetrahydrocannabinol, tetrahydocannabinolic acid A, cannabidiol, cannabidiolic acid, cannabigerol, cannabigerolic acid, cannabinol, cannabinolic acid, cannabichromene, cannabichromenic acid, cannabicyclol, cannabicyclolic acid, tetrahydrocannabivarin, tetrahydrocannabivarinic acid, cannabidivarin and cannabidivarinic acid. Resultant cannabinoid profiles were compared, and markers were suggested. Characteristics of Sativex® included a specific cannabidiol/tetrahydrocannabinol ratio and presence of cannabichromene, while acidic cannabinoids, cannabigerol and cannabinol occurred in only low amounts. As expected, the predominant ingredient in Dronabinol was tetrahydrocannabinol, but minor cannabinoids were quantified as well. Medical marihuana and seized cannabis were compared separately in a principal component analysis. Several medical marihuana varieties were found to significantly differ from seized cannabis, mostly regarding contents of tetrahydocannabinolic acid A and tetrahydrocannabivarinic acid and cannabidiolic and cannabidivarinic acid respectively., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Elsevier B.V. All rights reserved.)

The use of cannabidiol (CBD) in electronic cigarettes is widespread. Previously, it was reported that CBD is partly transformed to THC in case smoking as a cigarette, however, the pyrolysis of this compound has not been assessed extensively. The aim of our study was to investigate the effect of temperature on the composition of pyrolysis products of CBD. The experiments were performed in the typical operating temperature range of e-cigarettes (250-400 °C) and at 500 °C under both inert and oxidative conditions, and the pyrolysis products were identified and quantified by GC-MS. Depending on the temperature and atmosphere, 25-52% of CBD was transformed into other chemical substances: Δ 9 -THC, Δ 8 -THC, cannabinol and cannabichromene were the predominant pyrolysates in both conditions, all formed by cyclization reaction. THC was the main pyrolysis product at all temperatures under both oxidative and inert conditions. Our results point out that CBD in e-cigarettes can be considered as a precursor of THC, thus it bears all the dangers related to this psychoactive compound. Our findings are fundamental contributions to the safety profile of CBD-based e-cigarettes.

Stem cell therapy promotes tissue regeneration and wound healing. Efforts have been made to prime stem cells to enhance their regenerative abilities. Certain marijuana components, namely the non-psychoactive cannabidiol (CBD) and psychoactive tetrahydrocannabinol (THC), are defined as immunomodulators. 9 We test whether two sources of stem cells, primed with CBD or THC, would demonstrate improved regenerative abilities. Human adipose-derived stem cells (ASCs) and bone marrow-derived stem cells (BMDSCs), not obtained from the same individual, were treated with low (300 nM) or high (3 μM) concentration CBD. Porcine ASCs and BMDSCs were isolated from a single pig, and treated with either low or high concentrations of CBD or THC. Transwell migration and MTT proliferation assays were performed on the human ASCs and BMDSCs. Also, transwell migration assay was performed on the porcine ASCs and BMDSCs. Finally, a wound healing scratch assay in porcine primary fibroblasts (PFs) was performed, co-cultured with the cannabinoid-treated ASCs. CBD priming at low concentration induces migration by 180% (P < .01) in porcine ASCs, and by only 93% (P < .02) in porcine BMDSCs. In porcine stem cells, THC priming at low concentration induces migration by 91.6% (P < .01) in ASCs but by only 44.3% (P < .03) in BMDSCs. Compared to PFs co-cultured with untreated ASCs, PFs co-cultured with low CBD-primed ASCs had 75% faster wound closure at 18 hours (P < .01). CBD and THC priming of ASCs and BMDSCs, particularly at lower doses, enhances a number of regenerative parameters, suggesting that these major marijuana components may improve stem cell-based therapies. SIGNIFICANCE OF THE STUDY: Our study demonstrates that cannabinoids can enhance the regenerative capacity of two major sources of stem cells, adipose- and bone marrow-derived, from human and porcine donors. Stem cell isolation and expansion is invasive, costly and time consuming. Stem cells with improved regenerative properties may be effective in the treatment of acute or chronic wounds. This is the first study to compare the priming potential of two sources of stem cells from the same animal, with the same genetic and epigenetic profile, as well as the first to prime with THC., (© 2020 John Wiley & Sons Ltd.)

Advances in drug vapor exposure systems have enabled evaluation of Δ-9-tetrahydrocannabinol (THC) vapor effects in laboratory animals. The purpose of this study was to 1) establish a range of parameters of THC vapor exposure in rats sufficient to produce a behavioral dose-effect curve in a battery of tasks sensitive to THC; and 2) to investigate sex differences in the effects of THC vapor exposure and THC injection (intraperitoneal, IP) on these behaviors in two strains of outbred rats. Male and female Sprague Dawley and Wistar rats (N = 22, 5-6/sex per group) received THC via passive vapor exposure (200 mg/mL; 5 conditions) and IP injection (1-20 mg/kg) in a within subject design. The effects of vaped and injected THC on appetite was determined using progressive ratio responding for food pellets. THC effects on nociception, measured using the tail withdrawal assay, and body temperature were also assessed during a 5-h test period for evaluation of time course of effects. Plasma THC concentrations were assessed after THC vapor and 10 mg/kg IP THC. THC vapor produced exposure-related increases and decreases in motivation to obtain food under the progressive ratio schedule. IP THC (3-20 mg/kg) reduced breakpoints. Vaped and injected THC produced exposure and dose-dependent antinociception and hypothermia. Sex and strain differences in THC effects were also observed. Plasma THC concentrations were higher after 10 mg/kg IP THC (152 ng/mL) compared to the highest vapor exposure condition tested (38 ng/mL), but magnitude of behavioral effects were comparable. THC vapor exposure produced reliable, dose orderly effects on food-maintained behavior, nociception, and body temperature that are comparable to effects of IP THC, although there were differences in the time course of behavioral outcomes., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Introduction: Cannabidiol (CBD), the nonintoxicating constituent of cannabis, is largely employed for pharmaceutical and cosmetic purposes. CBD can be extracted from the plant or chemically synthesized. Impurities of psychotropic cannabinoids Δ 9 -tetrahydrocannabinol (Δ 9 -THC) and Δ 8 -THC have been found in extracted CBD, thus hypothesizing a possible contamination from the plant. Materials and Methods: In this study, synthetic and extracted CBD samples were analyzed by ultrahigh-performance liquid chromatography coupled to high-resolution mass spectrometry and the parameters that can be responsible of the conversion of CBD into THC were evaluated by an accelerated stability test. Results: In synthetic and extracted CBD no trace of THC species was detected. In contrast, CBD samples stored in the dark at room temperature on the benchtop for 3 months showed the presence of such impurities. Experiments carried out under inert atmosphere in the absence of humidity or carbon dioxide led to no trace of THC over time even at high temperature. Conclusions: The results suggested that the copresence of carbon dioxide and water from the air could be the key for creating the acidic environment responsible for the cyclization of CBD. These findings suggest that it might be appropriate to review the storage conditions indicated on the label of commercially available CBD., Competing Interests: No competing financial interests exist., (Copyright 2021, Mary Ann Liebert, Inc., publishers.)

The regulation of cannabinoid synthesis in Cannabis sativa is of increasing research interest as restrictions around the globe loosen to allow the plant's legal cultivation. Of the major cannabinoids, the regulation of cannabigerolic acid (CBGA) production is the least understood. The purpose of this study was to elucidate the inheritance of CBGA dominance in C. sativa and describe a marker related to this chemotype. We produced two crossing populations, one between a CBGA dominant cultivar and a tetrahydrocannabinolic acid (THCA) dominant cultivar, and one between a CBGA dominant cultivar and a cannabidiolic acid (CBDA) cultivar. Chemical and genotyping analyses confirmed that CBGA dominance is inherited as a single recessive gene, potentially governed by a non-functioning allelic variant of the THCA synthase. The "null" THCAS synthase contains a single nucleotide polymorphism (SNP) that may render the synthase unable to convert CBGA to THCA leading to the accumulation of CBGA. This SNP can be reliably used as a molecular marker for CBGA dominance in the selection and breeding of C. sativa .