Structure-based identification of a G protein-biased allosteric modulator of cannabinoid receptor CB1.

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 ß-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.

Potent, Selective Agonists for the Cannabinoid-like Orphan G Protein-Coupled Receptor GPR18: A Promising Drug Target for Cancer and Immunity.

The human orphan G protein-coupled receptor GPR18, activated by Δ9-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 ß-arrestin recruitment assays. Structure-activity relationships were analyzed, and selectivity versus cannabinoid (CB) and CB-like receptors was assessed. Compound 51 (PSB-KK1415, EC50 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, EC50 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.

Photodegradation of cannabidiol (CBD) and Δ9-THC in cannabis plant material.

Δ9-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 Δ9-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 Δ9-THC in three cannabis chemotypes, namely I (Δ9-THC 2.50%w/w), II (CBD:Δ9-THC 5.82%w/w:3.19%w/w) and III (CBD 3.02%w/w).

Differentiation of Δ9-THC and CBD Using Silver-Ligand Ion Complexation and Electrospray Ionization Tandem Mass Spectrometry (ESI-MS/MS).

The 2018 Farm Bill defines marijuana as Cannabis sativa L. or any derivative thereof that contains greater than 0.3% Δ9-tetrahydrocannabinol (Δ9-THC) on a dry weight basis. The main cannabinoids present in Cannabis sativa L., Δ9-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 Δ9-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 Δ9-THC and CBD using six different Ag complexes. Differences between the binding affinities of Δ9-THC and CBD for [Ag(PPh3)(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 Δ9-THC. When applied to the analysis of known Δ9-THC:CBD mixture ratios, the developed [Ag(PPh3)(OTf)]2 ion complexation method was able to differentiate Δ9-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 Δ9-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 Δ9-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.

Investigation of the intrinsic cannabinoid activity of hemp-derived and semisynthetic cannabinoids with ß-arrestin2 recruitment assays-and how this matters for the harm potential of seized drugs.

Cultivation of industrial low-Δ9-tetrahydrocannabinol (Δ9-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 Δ9-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 CB1 cannabinoid receptor with a ß-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 CB1 activation than Δ9-THC, based on either higher efficacy (Emax) or higher potency (EC50). Structure-activity relationships were assessed for Δ9-THC and Δ8-THC homologs encompassing elongated alkyl chains. Additionally, stereoisomer-specific differences in CB1 activity were established for various THC isomers (Δ7-THC, Δ10-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 Δ9-THC products.

A simple aptamer-dye fluorescence sensor for detecting Δ9-tetrahydrocannabinol and its metabolite in urban sewage.

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.

The Therapeutic Potential and Molecular Mechanisms Underlying the Neuroprotective Effects of Sativex® - A Cannabis-derived Spray.

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.

A forensic application of genetic markers for distinction between drug-type and fiber-type Cannabis sativa L.

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.

Preventing Mislabeling: A Comparative Chromatographic Analysis for Classifying Medical and Industrial Cannabis.

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 R2 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.

"Breaking bud": the effect of direct chemical modifications of phytocannabinoids on their bioavailability, physiological effects, and therapeutic potential.

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.

The B1080/B1192 molecular marker identifies hemp plants with functional THCA synthase and total THC content above legal limit.

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.

Delta-9-Tetrahydrocannabinolic Acid B: A Mechanism for its Formation in Cannabis.

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.

Cannabidiol and Δ8-Tetrahydrocannabinol: Cannabinoids of Rising Interest and Concern.

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.

E-cigarette vaping associated acute lung injury (EVALI): state of science and future research needs.

"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.

Elucidation of partial activation of cannabinoid receptor type 2 and identification of potential partial agonists: Molecular dynamics simulation and structure-based virtual screening.

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 Δ9-tetrahydrocannabinol (Δ9-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 Δ9-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 Δ9-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.

DoE-assisted development and validation of a stability-indicating HPLC-DAD method for simultaneous determination of five cannabinoids in Cannabis sativa L. based on analytical quality by design (AQbD) concept.

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, Δ9 -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 (PI95% ). 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.

Inhibition of human androgen receptor by delta 9-tetrahydro-cannabinol and cannabidiol related to reproductive dysfunction: A computational study.

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.

Vaping Aerosols from Vitamin E Acetate and Tetrahydrocannabinol Oil: Chemistry and Composition.

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 Δ9-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.

Mechanistic origin of partial agonism of tetrahydrocannabinol for cannabinoid receptors.

Cannabinoid receptor 1 (CB1) is a therapeutically relevant drug target for controlling pain, obesity, and other central nervous system disorders. However, full agonists and antagonists of CB1 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 Δ9-Tetrahydrocannabinol (THC) against CB1. 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 CB1 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.

LC-MS/MS Assay for the Measurement of Cannabidiol Profiling in CBD Oil from Japanese Market and Application for Convertible Tetrahydrocannabinol in Acetic Acid Condition.

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 C18 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.