Characterisation of Cannabis-Based Products Marketed for Medical and Non-Medical Use Purchased in Portugal.

Cannabis-based products have gained attention in recent years for their perceived therapeutic benefits (with cannabinoids such as THC and CBD) and widespread availability. However, these products often lack accurate labelling regarding their cannabinoid content. Our study, conducted with products available in Portugal, revealed significant discrepancies between label claims and actual cannabinoid compositions. A fully validated method was developed for the characterisation of different products acquired from pharmacies and street shops (beverages, herbal samples, oils, and cosmetic products) using high-performance liquid chromatography coupled with a diode array detector. Linearity ranged from 0.4 to 100 µg/mL (0.04-10 µg/mg) (THC, 8-THC, CBD, CBG, CBDA, CBGA), 0.1-100 µg/mL (0.01-10 µg/mg) (CBN), 0.4-250 µg/mL (0.04-25 µg/mg) (THCA-A), and 0.8-100 µg/mL (0.08-10 µg/mg) (CBCA). Among sampled beverages, none contained detectable cannabinoids, despite suggestive packaging. Similarly, oils often differed from the declared cannabinoid compositions, with some containing significantly higher CBD concentrations than labelled. These inconsistencies raise serious concerns regarding consumer safety and informed decision-making. Moreover, our findings underscore the need for stringent regulation and standardised testing protocols to ensure the accuracy and safety of cannabis-based products.

Ultra-high-performance liquid chromatography-tandem mass spectrometry assay for quantifying THC, CBD and their metabolites in hair. Application to patients treated with medical cannabis.

Recently, several countries approved the use of cannabis flowering tops with standardized amount of ∆9-tetrahydrocannabinol (THC), cannabidiol (CBD) to treat several diseases. Therapeutic monitoring of medical cannabis products administered to patients for the established pathologies is rarely carried out. Previous few investigations have been developed in conventional matrices like blood and urine. This is the first study involving hair analysis of THC, CBD and their metabolites in patients treated with medical cannabis. An ultra-high-performance liquid chromatography-tandem mass spectrometry method to quantify THC, CBD, and metabolites, i.e., 11-nor-9-carboxy-THC (THC-COOH), 11-hydroxy-THC (11-OH-THC) cannabidiol-7-oic acid (7-COOH-CBD), 7-hydroxycannabidiol (7-OH-CBD), 6-α-hydroxycannabidiol (6-α-OH-CBD) and 6-ß-hydroxycannabidiol (6-ß-OH-CBD) in hair samples was developed and fully validated. The validation results indicated that the method was accurate (average inter/intra-day error, <10%), precise (inter/intra-day imprecision, <10%), and fast (10 min run time). Average hair concentrations in four patients treated with different formulations of medical cannabis were 2.75 ng/mg THC, 2.87 ng/mg 11-OH-THC, and 0.32 ng/mg THC-COOH (n = 3); 1.65 ng/mg CBD, 2.73 ng/mg 7-OH-CBD, 1.29 ng/mg 7-COOH-CBD, 0.35 ng/mg 6-α-OH-CBD, and 0.03 ng/mg 6-ß-OH-CBD. The proposed method proved suitable for a fast and sensitive determination of all target compounds allowing high throughput testing in individuals monitored for medical cannabis treatments.

Impact of Lipid Sources on Quality Traits of Medical Cannabis-Based Oil Preparations.

The feasibility of the use of two lipid sources and their impact on the cannabinoid profile, terpene fingerprint, and degradation products in medical cannabis oil preparations during 3 months of refrigerated storage time were investigated. LCHRMS-Orbitrap® and HS-SPME coupled to GC-MS for the investigation of targeted and untargeted cannabinoids, terpenes, and lipid degradation products in Bedrocan® and Bediol® macerated oils were used as analytical approaches. As regards the cannabinoid trend during 90 days of storage, there were no differences between PhEur-grade olive oil (OOPH) and medium-chain triglycerides oil (MCT oil) coupled to a good stability of preparations for the first 60 days both in Bedrocan® and Bediol® oils. MCT lipid source extracted a significant concentration of terpenes compared to olive oil. Terpenes showed a different scenario since MCT oil displayed the strongest extraction capacity and conservation trend of all compounds during the shelf life. Terpenes remained stable throughout the entire storage period in MCT formulations while a significant decrease after 15 and 30 days in Bediol® and Bedrocan® was observed in olive oil. Therefore, MCT oil could be considered a more suitable lipid source compared to olive oil involved in the extraction of medical cannabis for magistral preparations.

Discrimination of legal and illegal Cannabis spp. according to European legislation using near infrared spectroscopy and chemometrics.

Aerial parts containing cannabidiol can be purchased in a legal way but cannabis used as recreational drug is illegal in most European countries. Δ9-tetrahydrocannabinol is one of the main cannabinoids responsible for the psychotropic effect. European Union countries and Switzerland authorize a concentration of THC of 0.2 % and 1.0 % w/w, respectively, for smoking products and industrial hemp. Public health inspectors and law enforcement officers need to check the legality of samples. Therefore there is a need for innovative approaches, allowing quality control of these products in an easy way and preferably on site. In many countries, cultivation of industrial hemp is permitted if the THC content does not exceed 0.2 % w/w. A portable equipment could be a useful measuring tool for farmers to check for the THC content at regular time. In this work, 189 samples were analysed with a benchtop and a handheld NIR device in order to create two classification methods according to European and Swiss laws. All samples were also analysed by GC-FID to determine their THC concentration. Supervised analysis was applied in order to establish the best model. For the first classification, the accuracy was 91% for the test set with the benchtop data and 93 % for the test set with the handheld data. For the second classification, the accuracies were respectively 91 % and 95 %. The obtained models, hyphenating spectroscopic techniques and chemometrics, enable to discriminate legal and illegal cannabis samples according to European and Swiss laws.

Fast and sensitive UHPLC-MS/MS analysis of cannabinoids and their acid precursors in pharmaceutical preparations of medical cannabis and their metabolites in conventional and non-conventional biological matrices of treated individual.

Monitoring pharmacological active compounds in pharmaceutical preparations of medical cannabis and in conventional and non-conventional biological matrices of treated individuals use requires both a wide linear range and sensitive detection. We have developed and validated a fast and sensitive method using ultra-high performance liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS/MS) for analysis of Δ-9-tetrahydrocannabinol (THC), cannabidiol (CBD), their acidic precursors Δ-9-tetrahydrocannabinolic acid A (THCA-A) and cannabidiolic acid (CBDA) and some major metabolites of THC such as 11-nor-9-carboxy-THC (THC-COOH), 11-hydroxy-THC (11-OH-THC), Δ-9-THC-Glucuronide (THC-GLUC) and THC-COOH-Glucuronide (THC-COOH-GLUC) in conventional (whole blood and urine) and non-conventional (oral fluid and sweat) of individual treated with medical cannabis preparation. Specifically, THC, THCA-A, CBD and CBD-A were determined in cannabis decoction and oil prepared to treat individuals. The method used positive electrospray ionization (ESI) mode to reach the sensitivity needed to detect minimal amounts of analytes under investigations exposure with limits of quantification ranging from 0.2 to 0.5 ng per milliliter (ng/mL) or ng per patch in case of collected sweat. The validation results indicated this method was accurate (average inter/intra-day error, <10%), precise (inter/intra-day imprecision, <10%), and fast (10 min run time). In addition, time-consuming sample preparation was avoided applying dilute and shoot procedure, meeting the needs for potential large-scale population studies. The analysis of real samples demonstrated a pharmacokinetics of cannabinoids, their precursors and their metabolites dependent from quantity of carboxylated and decarboxylated compounds in pharmaceutical preparations.

Generation of a Comprehensive Transcriptome Atlas and Transcriptome Dynamics in Medicinal Cannabis.

Cannabinoids are the main medicinal compounds of interest in the plant Cannabis sativa, that are primarily synthesised in the glandular trichomes; found on female floral buds. The content, composition and yield of secondary metabolites (cannabinoids and terpenoids) is influenced by the plant's genetics and environment. Some initial gene expression experiments have been performed from strains of this plant species that contrasted in cannabinoid production, however the present knowledge about detailed trichome transcriptomics in this species is limited. An extensive transcriptome atlas was generated by RNA sequencing using root, shoot, flower and trichome tissues from a female plant strain (Cannbio-2) and was enhanced with the addition of vegetative and reproductive tissues from a male cannabis plant. Differential gene expression analysis identified genes preferentially expressed in different tissues. Detailed trichomics was performed from extractions specifically from glandular trichomes as well as female floral tissues at varying developmental stages, to identify stage-specific differentially expressed genes. Candidate genes involved in terpene and cannabinoid synthesis were identified and the majority were found to have an abundant expression in trichomes. The comprehensive transcriptome is a significant resource in cannabis for further research of functional genomics to improve the yield of specialised metabolites with high pharmacological value.

Analytical quality by design: Development and control strategy for a LC method to evaluate the cannabinoids content in cannabis olive oil extracts.

Cannabidiol (CBD) and Δ9-tetrahydrocannabinol (Δ9-THC) are considered as the most interesting cannabinoids in Cannabis sativa L. for the clinical practice. Since 2013, the Italian law allows pharmacists to prepare and dispense cannabis extracts to patients under medical prescription, and requires the evaluation of CBD and Δ9-THC content in cannabis extracts before sale. Cannabis olive oil extracts are prepared from dried female cannabis inflorescences, but a standard protocol is still missing. In this study, a fast RP-HPLC/UV method has been developed to quantify CBD and Δ9-THC in cannabis olive oil extracts. The analytical quality by design strategy has been applied to the method development, setting critical resolution and total analysis time as critical method attributes (CMAs), and selecting column temperature, buffer pH and flow rate as critical method parameters. Information from Doehlert Design in response surface methodology combined to Monte-Carlo simulations led to draw the risk of failure maps and to identify the method operable design region. The method was validated according to the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH) guidelines and then implemented in routine analysis. A control strategy based on system control charts was planned to monitor the developed method performances. Evaluation data were recorded over a period of one year of routine use, and both the CMAs showed values within the specifications in every analysis performed. Hence, a new risk evaluation for the future performances of the method was achieved by using a Bayesian approach based on the routine use data, computing the future distribution of the two CMAs. Finally, a study focusing on the monitoring of CBD and Δ9-THC concentrations in cannabis olive oil extracts was carried out. The developed method was applied to 459 extracts. The statistical analysis of the obtained results highlighted a wide variability in terms of concentrations among different samples from the same starting typology of cannabis, underlining the compelling need of a standardised procedure to harmonise the preparation of the extracts.

Accumulation of bioactive metabolites in cultivated medical Cannabis.

There has been an increased use of medical Cannabis in the United States of America as more states legalize its use. Complete chemical analyses of this material can vary considerably between producers and is often not fully provided to consumers. As phytochemists in a state with legal medical Cannabis we sought to characterize the accumulation of phytochemicals in material grown by licensed commercial producers. We report the development of a simple extraction and analysis method, amenable to use by commercial laboratories for the detection and quantification of both cannabinoids and terpenoids. Through analysis of developing flowers on plants, we can identify sources of variability of floral metabolites due to flower maturity and position on the plant. The terpenoid composition varied by accession and was used to cluster cannabis strains into specific types. Inclusion of terpenoids with cannabinoids in the analysis of medical cannabis should be encouraged, as both of these classes of compounds could play a role in the beneficial medical effects of different cannabis strains.

Implementation of a generic SFC-MS method for the quality control of potentially counterfeited medicinal cannabis with synthetic cannabinoids.

In this study, we describe the development of a SFC-MS method for the quality control of cannabis plants that could be potentially adulterated with synthetic cannabinoids. Considering the high number of already available synthetic cannabinoids and the high rate of development of novel structures, we aimed to develop a generic method suitable for the analysis of a large panel of substances using seventeen synthetic cannabinoids from multiple classes as model compounds. Firstly, a suitable column was chosen after a screening phase. Secondly, optimal operating conditions were obtained following a robust optimization strategy based on a design of experiments and design space methodology (DoE-DS). Finally, the quantitative performances of the method were assessed with a validation according to the total error approach. The developed method has a run time of 9.4 min. It uses a simple modifier composition of methanol with 2% H2O and requires minimal sample preparation. It can chromatographically separate natural cannabinoids (except THC-A and CBD-A) from the synthetics assessed. Also, the use of mass spectrometry provides sensitivity and specificity. Moreover, this quality by design (QbD) approach permits the tuning of the method (within the DS) during routine analysis to achieve a desirable separation since the future compounds that should be analyzed could be unknown. The method was validated for the quantitation of a selected synthetic cannabinoid in fiber-type cannabis matrix over the range of 2.5% - 7.5% (w/w) with LOD value as low as 14.4 ng/mL. This generic method should be easy to implement in customs or QC laboratories in the context of counterfeit drugs tracking.

Multiresidue Method of Analysis of Pesticides in Medical Cannabis.

Three related analytical methods were developed and validated for the determination of pesticides in cannabis leaves, dried cannabis flowers, and cannabis oil. The methods follow the generic sequence of an acetonitrile extraction, followed by solid-phase extraction cleanup and analysis by HPLC-tandem mass spectrometry (HPLC-MS/MS), GC-MS/MS, and GC-MS. These methods were developed to accommodate sample quantity and lipid content of the different matrices. Validation at a spiking level of 0.01 µg/g was successful for 39 pesticides in cannabis leaves and 40 pesticides in cannabis oil, and at 0.02 µg/g for 32 pesticides in cannabis flowers, with the majority of analytes showing recoveries within the acceptable range of 70-130%. With these methods established, unannounced inspections of Canadian licensed producers of cannabis revealed that out of 144 samples collected, 26 showed the presence of unauthorized pest control products.

Cannabis for Chronic Pain: Challenges and Considerations.

The National Academies of Sciences, Engineering, and Medicine has found substantial evidence that cannabis (plant) is effective for the treatment of chronic pain in adults, and moderate evidence that oromucosal cannabinoids (extracts, especially nabiximols) improve short-term sleep disturbances in chronic pain. The paradoxical superiority of the cannabis plant over cannabinoid molecules represents a challenge for the medical community and the established processes that define modern pharmacy. The expanding and variable legalization of cannabis in multiple states nationwide represents an additional challenge for patients and the medical community because recreational and medicinal cannabis are irresponsibly overlapped. Cannabis designed for recreational use (containing high levels of active ingredients) is increasingly available to patients with chronic pain who do not find relief with current pharmacologic entities, which exposes patients to potential harm. This article analyzes the available scientific evidence to address controversial questions that the current state of cannabis poses for health care professionals and chronic pain patients and sets the basis for a more open discussion about the role of cannabis in modern medicine for pain management. A critical discussion on these points, the legal status of cannabis, and considerations for health care providers is presented.

Comprehensive quality evaluation of medical Cannabis sativa L. inflorescence and macerated oils based on HS-SPME coupled to GC-MS and LC-HRMS (q-exactive orbitrap®) approach.

There are at least 554 identified compounds in C. sativa L., among them 113 phytocannabinoids and 120 terpenes. Phytocomplex composition differences between the pharmaceutical properties of different medical cannabis chemotype have been attributed to strict interactions, defined as 'entourage effect', between cannabinoids and terpenes as a result of synergic action. The chemical complexity of its bioactive constituents highlight the need for standardised and well-defined analytical approaches able to characterise the plant chemotype, the herbal drug quality as well as to monitor the quality of pharmaceutical cannabis extracts and preparations. Hence, in the first part of this study an analytical procedures involving the combination of headspace-solid-phase microextraction (HS-SPME) coupled to GC-MS and High Resolution Mass-Spectrometry LC-HRMS (Orbitrap®) were set up, validated and applied for the in-depth profiling and fingerprinting of cannabinoids and terpenes in two authorised medical grade varieties of Cannabis sativa L. inflorescences (Bedrocan® and Bediol®) and in obtained macerated oils. To better understand the trend of all volatile compounds and cannabinoids during oil storage a new procedure for cannabis macerated oil preparation without any thermal step was tested and compared with the existing conventional methods to assess the potentially detrimental effect of heating on overall product quality.

Pharmaceutical and biomedical analysis of cannabinoids: A critical review.

Cannabis products have recently regained much attention due to the high pharmacological potential of their cannabinoid content. In this review, the most widely used sample preparation strategies for the extraction of cannabinoids are described for the specific application to either plant materials or biological matrices. Several analytical techniques are described pointing out their respective advantages and drawbacks. In particular, chromatographic methods, such as TLC, GC and HPLC, are discussed and compared in terms of selectivity and sensitivity. Various detection methods are also presented based on the specific aim of the cannabinoids analysis. Lastly, critical considerations are mentioned with the aim to deliver useful suggestions for the selection of the optimal and most suitable method of analysis of cannabinoids in either biomedical or cannabis derived samples.

Asociaciones cannábicas: una respuesta social para un reto global / No disponible

Se resumen los objetivos y las cinco propuestas (los cinco pilares) de los clubs sociales de cannabis, en el contexto del movimiento cannábico que busca promocionar nuevas políticas hacia la regulación de la producción, distribución y consumo de la planta, excluyendo su explotación comercial. También se reclama el acceso a la marihuana terapéutica y a la educación para un consumo responsable

The objectives and proposals (the five pillars) of the cannabis social clubs are summarized. Aim of the cannabis movement is to promote new policies for the regulation of production, distribution and use of the plant. Commercial exploitation of the crop is excluded. Access to therapeutic marijuana and education for a responsible use are also claimed

Medicinal cannabis: Principal cannabinoids concentration and their stability evaluated by a high performance liquid chromatography coupled to diode array and quadrupole time of flight mass spectrometry method.

In the last few years, there has been a boost in the use of cannabis-based extracts for medicinal purposes, although their preparation procedure has not been standardized but rather decided by the individual pharmacists. The present work describes the development of a simple and rapid high performance liquid chromatography method with UV detection (HPLC-UV) for the qualitative and quantitative determination of the principal cannabinoids (CBD-A, CBD, CBN, THC and THC-A) that could be applied to all cannabis-based medicinal extracts (CMEs) and easily performed by a pharmacist. In order to evaluate the identity and purity of the analytes, a high-resolution mass spectrometry (HPLC-ESI-QTOF) analysis was also carried out. Full method validation has been performed in terms of specificity, selectivity, linearity, recovery, dilution integrity and thermal stability. Moreover, the influence of the solvent (ethyl alcohol and olive oil) was evaluated on cannabinoids degradation rate. An alternative extraction method has then been proposed in order to preserve cannabis monoterpene component in final CMEs.

Medicinal Cannabis: In Vitro Validation of Vaporizers for the Smoke-Free Inhalation of Cannabis.

Inhalation by vaporization is a promising application mode for cannabis in medicine. An in vitro validation of 5 commercial vaporizers was performed with THC-type and CBD-type cannabis. Gas chromatography/mass spectrometry was used to determine recoveries of total THC (THCtot) and total CBD (CBDtot) in the vapor. High-performance liquid chromatography with photodiode array detection was used for the quantitation of acidic cannabinoids in the residue and to calculate decarboxylation efficiencies. Recoveries of THCtot and CBDtot in the vapor of 4 electrically-driven vaporizers were 58.4 and 51.4%, 66.8 and 56.1%, 82.7 and 70.0% and 54.6 and 56.7% for Volcano Medic®, Plenty Vaporizer®, Arizer Solo® and DaVinci Vaporizer®, respectively. Decarboxylation efficiency was excellent for THC (≥ 97.3%) and CBD (≥ 94.6%). The gas-powered Vape-or-Smoke™ showed recoveries of THCtot and CBDtot in the vapor of 55.9 and 45.9%, respectively, and a decarboxylation efficiency of ≥ 87.7 for both cannabinoids. However, combustion of cannabis was observed with this device. Temperature-controlled, electrically-driven vaporizers efficiently decarboxylate inactive acidic cannabinoids and reliably release their corresponding neutral, active cannabinoids. Thus, they offer a promising application mode for the safe and efficient administration of medicinal cannabis.

Understanding dabs: contamination concerns of cannabis concentrates and cannabinoid transfer during the act of dabbing.

Cannabis concentrates are gaining rapid popularity in the California medical cannabis market. These extracts are increasingly being consumed via a new inhalation method called 'dabbing'. The act of consuming one dose is colloquially referred to as "doing a dab". This paper investigates cannabinoid transfer efficiency, chemical composition and contamination of concentrated cannabis extracts used for dabbing. The studied concentrates represent material available in the California medical cannabis market. Fifty seven (57) concentrate samples were screened for cannabinoid content and the presence of residual solvents or pesticides. Considerable residual solvent and pesticide contamination were found in these concentrates. Over 80% of the concentrate samples were contaminated in some form. THC max concentrations ranged from 23.7% to 75.9% with the exception of one outlier containing 2.7% THC and 47.7% CBD. Up to 40% of the theoretically available THC could be captured in the vapor stream of a dab during inhalation experiments. Dabbing offers immediate physiological relief to patients in need but may also be more prone to abuse by recreational users seeking a more rapid and intense physiological effect.