Identification of 5F-Cumyl-PINACA, a Synthetic Cannabinoid, in the Herbal Material Used for Recreational Purposes in the Province of Trieste: Public Health Implications.

BACKGROUND: In recent years, the phenomenon of the production and trade of synthetic cannabinoids has grown, becoming a public health issue worldwide. The recent access- to the ED of the hospital of Trieste- of people who complained of episodes of hallucinations, sensation of poisoning, tachycardia, and air hunger following the inhalation of "Che Sballo platinum", have highlighted the need to perform further analysis on the contents of the packet sold as an air freshener, produced in Koper (Slovenia). OBJECTIVE: This paper wants to be an alert about the possible consequences on health due to the spreading of "Che Sballo platinum" in the province of Trieste. METHODS: The package contents were analyzed by a multi-target screening method of MRM-IDAEPI experiment. The result was then confirmed, and quantification was achieved via LC-ESI-MS/MS analysis in MRM mode using QTrap 6500 + Sinergy hydro column 100 x 2 mm 1.9 um transitions MRM1 368.3 → 250.0; MRM2 368.3 → 233.0. RESULTS: The initial screening tested negative for THC and showed positive results for 5F-Cumyl- PINACA. Quantitation result reported dose by the package of 8.5 mg of the compound. Formal notification was sent to the Italian Health Authorities (Notification No. 2021110205). CONCLUSION: Consumption of 5F-Cumyl-PINACA results in much more potent effects than marijuana. Lack of information about the actual concentration of the substance on the packaging does not allow drug users to have an adequate dosage, with possible toxic consequences on health. Further investigations must be done to discover the true extent of the phenomenon.

Prevalence of cathinones and other new psychoactive substances in hair of parents and children of families with known or suspected parental abuse of conventional illegal drugs.

BACKGROUND: Hair analysis of parents and their children was regularly used since 2011 as a diagnostic tool in a social support project for families with known or suspected abuse of conventional illegal drugs and revealed a high incidence of cocaine, cannabinoids, amphetamines, ecstasy and heroin. In this context, the prevalence of new psychoactive substances (NPS) in these families should be important for a realistic estimate of the situation. METHODS: The extracts of 1537 hair samples from 318 children (age 1-14 years), 44 adolescents and 611 adults, which were collected and tested for conventional drugs between June 2016 and April 2021 and frozen at -20 °C, were reanalyzed by a validated LC-MS/MS method (limits of quantitation 5-24 pg/mg) for 33 cathinones, 10 phenylethylamines, 5 piperazines including the antidepressant trazodone, 2 tryptamines, 9 designer benzodiazepines, 4 synthetic opioids and 4 ketamine-like substances including phencyclidine. RESULTS: Between one and up to five from 42 of these substances were detected in 227 samples (14.8%). The most frequently detected substances were benzedrone (62x), α-pyrrolidinovalerophenone (41x), N-ethylamphetamine (29x), dimethyltryptamine (13x) and pyrovalerone (11x). The quantification was possible only for 34 results of 15 drugs and the remaining majority of the results were unambiguously identified below LLOQ. The relative frequency of conventional drugs in the 227 NPS positive samples was higher than in all 1310 NPS negative samples for cocaine (69.6% vs. 56.0%), heroin (6-acetylmorphine 8.8% vs. 4.9%), amphetamine (16.3% vs. 7.7%) and MDMA (16.3% vs. 7.0%) but was similar for THC (38.3% vs. 36.3%) and benzodiazepines (1.8% vs. 1.7%). The high prevalence of N-ethylamphetamine can be explained as a byproduct of the illicit amphetamine synthesis from benzaldehyde and nitroethane rather than as a separate drug or as a combined metabolite of amphetamine and ethanol. The isolated appearance of 3-trifluoromethylphenylpiperazine in 9 hair samples collected in January 2017 can be caused either by its use as an NPS or by its formation as a metabolite of the medical drug flibanserin. The results were compared within 17 families whose members were tested at the same time and showed positive NPS results. The detected drugs agreed between both parents only in about half of the cases whereas the drugs found in children's hair was always detected also in hair of one or both parents. CONCLUSION: The re-testing of hair extracts for NPS after long-time storage in frozen state enables an impression about the relative high prevalence in the tested population group, despite the limitation by partial degradation of the substances and the corresponding impossibility in quantitative assessments. In addition to conventional drugs, the hair test for these substances should be useful in unclear cases of child's welfare endangerment and in family law.

Direct and Fast Screening of New Psychoactive Substances Using Medical Swabs and Atmospheric Solids Analysis Probe Triple Quadrupole with Data-Dependent Acquisition.

New psychoactive substances (NPS) have become a serious public health problem, as they are continuously changing their structures and modifying their potency and effects on humans, and therefore, novel compounds are unceasingly appearing. One of the major challenges in forensic analysis, particularly related to the problem of NPS, is the development of fast screening methodologies that allow the detection of a wide variety of compounds in a single analysis. In this study, a novel application of the atmospheric solids analysis probe (ASAP) using medical swabs has been developed. The swab-ASAP was coupled to a triple quadrupole mass analyzer working under a data-dependent acquisition mode in order to perform a suspect screening of NPS in different types of samples as well as on surfaces. The compounds were automatically identified based on the observed fragmentation spectra using an in-house built MS/MS spectra library. The developed methodology was applied for the identification of psychoactive substances in research chemicals and herbal blends. The sensitivity of the method, as well as its applicability for surface analysis, was also assessed by identifying down to 1 µg of compound impregnated onto a laboratory table. Another remarkable application was the identification of cathinones and synthetic cannabinoids on the fingers of potential consumers. Interestingly, our data showed that NPS could be identified on the fingers after being in contact with the product and even after cleaning their hands by shaking off with a cloth. The methodology proposed in this paper can be applied for routine analyses of NPS in different matrix samples without the need to establish a list of target compounds prior to analysis.

Pharmacological and biotransformation studies of 1-acyl-substituted derivatives of d-lysergic acid diethylamide (LSD).

The ergoline d-lysergic acid diethylamide (LSD) is one of the most potent psychedelic drugs. 1-Acetyl-LSD (ALD-52), a derivative of LSD containing an acetyl group on the indole nitrogen, also produces psychedelic effects in humans and has about the same potency as LSD. Recently, several other 1-acyl-substitued LSD derivatives, including 1-propanoyl-LSD (1P-LSD) and 1-butanoyl-LSD (1B-LSD), have appeared as designer drugs. Although these compounds are assumed to act as prodrugs for LSD, studies have not specifically tested this prediction. The present investigation was conducted to address the gap of information about the pharmacological effects and mechanism-of-action of 1-acyl-substituted LSD derivatives. Competitive binding studies and calcium mobilization assays were performed to assess the interaction of ALD-52, 1P-LSD, and 1B-LSD with serotonin 5-HT2 receptor subtypes. A receptorome screening was performed with 1B-LSD to assess its binding to other potential targets. Head twitch response (HTR) studies were performed in C57BL/6J mice to assess in vivo activation of 5-HT2A (the receptor thought to be primarily responsible for hallucinogenesis). Finally, liquid chromatography/ion-trap mass spectrometry (LC/MS) was used to quantify plasma levels of LSD in Sprague-Dawley rats treated with ALD-52 and 1P-LSD. 1-Acyl-substitution reduced the affinity of LSD for most monoamine receptors, including 5-HT2A sites, by one to two orders of magnitude. Although LSD acts as an agonist at 5-HT2 subtypes, ALD-52, 1P-LSD and 1B-LSD have weak efficacy or act as antagonists in Ca2+-mobilization assays. Despite the detrimental effect of 1-acyl substitution on 5-HT2A affinity and efficacy, 1-acyl-substitued LSD derivatives induce head twitches in mice with relatively high potency. High levels of LSD were detected in the plasma of rats after subcutaneous administration of ALD-52 and 1P-LSD, demonstrating these compounds are rapidly and efficiently deacylated in vivo. These findings are consistent with the prediction that ALD-52, 1P-LSD and 1B-LSD serve as prodrugs for LSD. This article is part of the special issue entitled 'Serotonin Research: Crossing Scales and Boundaries'.

Biomedical analysis of New Psychoactive Substances (NPS) of natural origin.

New psychoactive substances (NPS) can be divided into two main groups: synthetic molecules and active principles of natural origin. With respect to this latter group, a wide range of alkaloids contained in plants, mainly from Asia and South America, can be included in the class of NPS of natural origin. The majority NPS of natural origin presents stimulant and/or hallucinogenic effects (e.g. Catha edulis and Ayahuasca, respectively) while few of them show sedative and relaxing properties (e.g. kratom). Few information is available in relation to the analytical identification of psychoactive principles contained in the plant material. Moreover, to our knowledge, scarce data are present in literature, about the characterization and quantification of the parent drug in biological matrices from intoxication and fatality cases. In addition, the metabolism of natural active principles has not been yet fully investigated for most of the psychoactive substances from plant material. Consequently, their identification is not frequently performed and produced metabolites are often unknown. To fill this gap, we reviewed the currently available analytical methodologies for the identification and quantification of NPS of natural origin in plant material and, whenever possible, in conventional and non-conventional biological matrices of intoxicated and dead subjects. The psychoactive principles contained in the following plants were investigated: Areca catechu, Argyreia nervosa, Ayahuasca, Catha edulis, Ipomoea violacea, Mandragora officinarum, Mitragyna speciosa, Pausinystalia yohimbe, Piper methisticum, Psilocybe, Rivea corymbosa, Salvia divinorum, Sceletium tortuosum, Lactuca virosa. From the results obtained, it can be evidenced that although several analytical methods for the simultaneous quantification of different molecules from the same plants have been developed and validated, a comprehensive method to detect active compounds from different natural specimens both in biological and non-biological matrices is still lacking.

Cardiotoxicity screening of illicit drugs and new psychoactive substances (NPS) in human iPSC-derived cardiomyocytes using microelectrode array (MEA) recordings.

The use of recreational drugs, including new psychoactive substances (NPS), is paralleled by emergency department visits of drug users with severe cardiotoxicity. Drug-induced cardiotoxicity can be the (secondary) result of increased norepinephrine blood concentrations, but data on potential drug-induced direct effects on cardiomyocyte function are scarce. The presence of hundreds of NPS therefore calls for efficient screening models to assess direct cardiotoxicity. We investigated effects of four reference compounds (3-30 nM dofetilide, nifedipine and isoproterenol, and 1-10 µM mexiletine) and six recreational drugs (0.01-100 µM cocaine, 0.01-1000 µM amphetamine, MDMA, 4-fluoroamphetamine, α-PVP and MDPV) on cardiomyocyte function (beat rate, spike amplitude and field potential duration (FPD ≈ QT interval in ECGs)), using Pluricyte® human-induced pluripotent stem cell (hiPSC)-derived cardiomyocytes cultured on ready-to-use CardioPlate™ multi-well microelectrode arrays (mwMEAs). Moreover, the effects of exposure to recreational drugs on cell viability were assessed. Effects of reference compounds were in accordance with the literature, indicating the presence of hERG potassium (dofetilide), sodium (mexiletine) and calcium (nifedipine) channels and α-adrenergic receptors (isoproterenol). All recreational drugs decreased the spike amplitude at 10-100 µM. All amphetamine-type stimulants and α-PVP decreased the beat rate at 300 µM, while cocaine and MDPV did so at 10 µM and 30 µM, respectively. All drugs increased the FPD, however at varying concentrations. MDMA, MDPV and amphetamine affected cardiomyocyte function at concentrations relevant for human exposure, while other drugs affected cardiomyocyte function only at higher concentrations (≥ 10 µM). Cell viability was only mildly affected at concentrations well above the lowest concentrations affecting cardiomyocyte function. We demonstrate that MEA recordings of hiPSC-derived cardiomyocytes enable screening for acute, direct effects on cardiomyocyte function. Our data further indicate that tachycardia in patients exposed to recreational drugs is likely due to indirect drug effects, while prolonged repolarization periods (prolonged QTc interval) could (partly) result from direct drug effects on cardiomyocyte function.

Ion mobility spectrometry as a fast screening tool for synthetic cannabinoids to uncover drug trafficking in jail via herbal mixtures, paper, food, and cosmetics.

The greatest challenge for European drug policies is how to effectively respond to the dynamic and constantly changing market for new psychoactive substances (NPS). Even small modifications in the chemical structure of substances often allow circumventing existing laws. Also in prison, the consumption of NPS is rising and there is growing evidence that NPS are responsible for a large share of drug-related problems. Ion mobility spectrometry (IMS) is the technique of choice for trace analysis of illicit drugs or explosives at security points, for example airports. Currently, databases of the reduced mobility (K0 ) values are limited to classical drugs and should be completed with data of emerging NPS. In this article, K0 values, LODs (0.7-3.6 ng) and drift times of 25 synthetic cannabinoids were evaluated. The data were added to existing databases of IMS which were then applied for fast screening in prison. The detection capability of the portable IMS technique was evaluated by the determination of intra-day (0.089%) and inter-day precision (0.004% to 0.14%), systematic error (0.19%), and separation capability for structurally related NPS. The applicability of the methodology was demonstrated by the successful analysis of 12 different pieces of paper impregnated with synthetic cannabinoids, 7 different cosmetics, and 5 food samples (liquids), spiked with a mixture of narcotic drugs and a synthetic cannabinoid. In addition, 14 herbal mixtures and 36 different casework samples from prisons were analyzed provided by the State Office of Criminal Investigation Rhineland-Palatinate (Germany).

Understanding and managing the new psychoactive substances phenomenon: a holistic approach.

The new psychoactive substances (NPS) phenomenon has emerged as a global threat that challenges public health and institutions. There are important qualitative differences between the NPS and traditional drugs phenomena. We discuss these differences and explore the complex structure of the NPS phenomenon. We analyse the entire phenomenon with a global, holistic approach. We present an original framework to help policy makers, healthcare practitioners, and community workers understand the NPS phenomenon's structure and to plan comprehensive policy responses and prevention strategies. We discuss fundamental characteristics, driving forces, routes of information, and social and individual health risks of the phenomenon. We conclude that a holistic approach integrating all aspects of the framework is essential for addressing this emerging threat. We give practical examples of interventions likely to be effective.

A case of synthetic cannabinoid poisoning in Croatia.

The number of new psychoactive substances (NPS), synthetic cannabinoids (SCs) in particular, is growing constantly. Because of the insufficiently explored effects on consumer health, they have become a major problem in the emergency departments. They are difficult to identify, and there are no antidotes that could reverse their detrimental effects. We report a case of poisoning of a young man who used SCs. The patient was admitted to the emergency department of the Clinical Hospital Merkur, Zagreb (Croatia) after sniffing and smoking a herbal product bought on the street. He presented with severe cognitive difficulties and visible eye redness. Other symptoms included somnolence, disorientation, loss of coordination, unsteady gait, hyporeflexia, stiffness, cramps and cold limbs, blurred vision, teeth grinding, dry mouth, tinnitus, fear, suicidal thoughts, impaired focus, memory, and speech, sedation, fatigue, depression, thought blocking, and autistic behaviour. His skin was dry, and his mucosa dry and irritated. Herbal products "Rainbow Special" and "Luminated Aroma" used by the patient were qualitatively analysed with gas chromatography / mass spectrometry (GC/MS) after direct extraction with an organic solvent. Solid-phase extraction method was used to analyse serum and urine samples. Despite the negative findings of biological samples, mostly due to the limitations of GC/MS, the clinical picture infallibly pointed to the poisoning with SCs. This was confirmed by the findings of 5-fluoro AMB (methyl 2-(1-(5-fluoropentyl)-1H-indazole-3-carboxamido)-3-methylbutanoate) in the herbal products.

Intended and unintended use of cathinone mixtures.

INTRODUCTION: Cathinones are one of the most popular categories of new psychoactive substances (NPS) consumed. Cathinones have different pharmacological activities and receptor selectivity for monoamine transporters based on their chemical structures. They are incorporated into NPS mixtures and used with other NPS or 'traditional' drugs. Cathinone use represents significant health risks to individuals and is a public health burden. METHODS: Evidence of poly-NPS use with cathinones, seizure information, and literature analyses results on NPS mixtures was systematically gathered from online database sources, including Google Scholar, Scopus, Bluelight, and Drugs-Forum. RESULTS AND DISCUSSION: Results highlight the prevalence of NPS with low purity, incorporation of cathinones into NPS mixtures since 2008, and multiple members of the cathinone family being present in individual UK-seized samples. Cathinones were identified as adulterants in NPS marketed as being pure NPS, drugs of abuse, branded products, herbal blends, and products labelled "not for human consumption." Toxicity resulting from cathinone mixtures is unpredictable because key attributes remain largely unknown. Symptoms of intoxication include neuro-psychological, psychiatric, and metabolic symptoms. Proposed treatment includes holistic approaches involving psychosocial, psychiatric and pharmacological interventions. CONCLUSION: Raising awareness of NPS, education, and training of health care professionals are paramount in reducing harms related to cathinone use.

Identification and determination of synthetic cannabinoids in herbal products by dry film attenuated total reflectance-infrared spectroscopy.

A new procedure has been developed for the identification and quantitative determination of synthetic cannabinoids in illicit herbal preparations. The methodology is based on the use of attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) measurement of sample extracts with 2-propanol drying 5µL of the extracts onto the ATR crystal. The qualitative identification was carried out on the 2-propanol extract after identification of the herbal matrix, followed by its subtraction and using a cut-off criterion of 75%. Quantitative determination was made by univariate calibration using the absorbance of the band located at 1520cm-1 of the spectrum. Four different cannabinoids, RCS-4, JWH-210, UR-144 and JWH-081 were used as test analytes and the ATR-FTIR method provided limits of quantification from 14 till 79mgL-1. Sized blank market samples were successfully identified and UR-144 quantified.

Pharmacology should be at the centre of all preclinical and clinical studies on new psychoactive substances (recreational drugs).

Despite the publication of a substantial body of preclinical and clinical information on recent recreational drugs such as 3,4-methylenedioxymethamphetamine (MDMA, 'ecstasy') and cathinone compounds such as mephedrone there remains a disturbing lack of consensus as to how dangerous these compounds are to the health of the individual and to society in general. This perspective proposes that use of good pharmacological practice should be mandatory in all preclinical and clinical studies. Its use will assist both translation and reverse translation of information produced in animals and clinical subjects. We propose several basic rules to be followed in all future studies. Preclinical studies should employ pharmacokinetic-pharmacodynamic integration thereby exposing animals to known or calculable drug concentrations. This will provide results relevant to pharmacology rather than toxicology and, crucially, data relevant to human drug use. Full experimental detail should be routinely provided, to allow comparison with other similar work. In clinical studies evidence should be provided that the drug under investigation has been ingested by the subjects being examined, and details given of all other drugs being ingested. Drug-drug interactions are an unavoidable confound but studies of a size that allows reliable statistical evaluation and preferably allows sub-group analysis, particularly by using meta-analysis, should help with this problem. This may require greater collaboration between investigative groups, as routinely occurs during pharmaceutical clinical trials. Other proposals include greater integration of preclinical and clinical scientists in both preclinical and clinical studies and changes in the law regarding Good Manufacturing Process (GMP) sourcing of drug for human studies.