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Recently, semi-synthetic cannabinoids have entered the illegal market and are produced to mimic the psychoactive effects of tetrahydrocannabinol (THC). This is a case report of a 19-year-old man, who was hospitalized due to severe sedation, hypotension and tachy- and bradycardia after ingestion of the semi-synthetic cannabinoids hexahydrocannabinol (HHC) and hexahydrocannabiphorol (HHC-P) mixed in food. HHC-P, HHC and metabolites were identified in blood samples.

Emergence of semi-synthetic cannabinoids in cannabis products seized in Eastern Denmark over a 6-year period.

Semi-synthetic cannabinoids (SSCs) are derivatives of phytocannabinoids with slight chemical modifications. SSCs have appeared as legal alternatives to tetrahydrocannabinol (Δ-THC) in recent years. This study investigates the prevalence of SSCs in seized drug samples from Danish police and custom authorities seized in Eastern Denmark in the period 2018-2023.

Overview of systematic toxicological analysis strategies and their coverage of substances in forensic toxicology.

Systematic toxicological analysis (STA) is the process of using an adequate analytical methodology to detect and identify as many potentially toxicologically relevant compounds as possible in biological samples. STA is an important part of everyday routine work within forensic toxicology, and several methods for STA have frequently been published and reviewed independently. However, the many drugs and other substances involved, as well as the constant emergence of new ones, may pose a major challenge in STA, which often demands a strategy involving multiple analytical methods in parallel.

Exploring Enzymatic Hydrolysis of Urine Samples for Investigation of Drugs Associated with Drug-Facilitated Sexual Assault.

Analyzing urine is common in drug-facilitated sexual assault cases if the analysis of blood is not optimal. The efficient enzymatic pretreatment of urine is important for cleaving glucuronides and improving the detection of the parent drug. The aim was to investigate the efficiency of three β-glucuronidases on eleven glucuronides relevant to DFSA at different incubation periods and temperatures.

Evidence of 11-Hydroxy-hexahydrocannabinol and 11-Nor-9-carboxy-hexahydrocannabinol as Novel Human Metabolites of Δ-Tetrahydrocannabinol.

(-)--Δ-tetrahydrocannabinol (Δ-THC) is the primary psychoactive compound in the Cannabis sativa plant. Δ-THC undergoes extensive metabolism, with the main human phase I metabolites being 11-hydroxy-tetrahydrocannabinol (11-OH-THC) and 11-nor-9-carboxy-tetrahydrocannabinol (THC-COOH). Early animal studies have indicated that the 9-10 double bond may be reduced in vivo to yield 11-hydroxy-hexahydrocannabinol (11-OH-HHC) and 11-nor-9-carboxy-hexahydrocannabinol (HHC-COOH).

Driving under the influence of nitrous oxide - A retrospective study of HS-GC-MS analysis in whole blood.

Since 2020, our lab has received blood samples from traffic cases involving suspicion of driving under the influence of nitrous oxide (NO). While NO analysis by gas chromatography (GC) has been around for decades, quantitative results in blood from drivers have been only scarcely reported. We present a three-year (2020-2022) retrospective study of NO from traffic cases in Eastern Denmark with suspected involvement of NO intake.

Suitability of cardiac blood, brain tissue, and muscle tissue as alternative matrices for toxicological evaluation in postmortem cases.

Drug concentrations in peripheral blood are often used to evaluate whether death was caused by drug intoxication. In some cases, peripheral blood is not available, and analytical results of alternative matrices should instead be used in the toxicological evaluation. However, reference concentrations of alternative matrices are few, which makes interpretation of results a challenge.

Driving under the Influence of Amphetamine: Analytical Evaluation of Illegal or Prescription Drug Intake Using Chiral UHPLC-MS-MS.

Differentiation between consumption of illegal and prescription drugs remains an important aspect in forensic toxicology. While illicit amphetamine is most often racemic, the medicinal drugs marketed in Denmark for the treatment of attention-deficit hyperactivity disorder contain the pure (S)-enantiomer or a prodrug thereof. In this study, we present a simple and efficient analytical workflow to provide information about the origin of amphetamine consumed in forensic cases concerning driving under the influence of drugs (DUID).

Analytical reproducibility evaluated from duplicate measurements of authentic ante- and postmortem blood samples by LC-MS/MS compared with long-term imprecision estimates from quality control samples.

Precision (or imprecision) is one of the central performance parameters of all analytical methods. It is often evaluated during method validation in spiked samples, with a relatively low number of measurements typically during a short period. Validation results are well documented in the literature; however, evaluation of imprecision in authentic cases compared with long-term imprecision from quality control samples has not often been reported on.

A New Strategy for Efficient Retrospective Data Analyses for Designer Benzodiazepines in Large LC-HRMS Datasets.

The expanding and dynamic market of new psychoactive substances (NPSs) poses challenges for laboratories worldwide. The retrospective data analysis (RDA) of previously analyzed samples for new targets can be used to investigate analytes missed in the first data analysis. However, RDA has historically been unsuitable for routine evaluation because reprocessing and reevaluating large numbers of forensic samples are highly work- and time-consuming.

Comparison of Comprehensive Screening Results in Postmortem Blood and Brain Tissue by UHPLC-QTOF-MS.

Alternative specimens collected during autopsies can be valuable in postmortem toxicology in cases where peripheral blood is not available. The applicability of brain tissue as an alternative matrix for drug screening by ultra-high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry was investigated in this study. Results of the 50 most frequently detected drugs and metabolites of toxicological interest in blood and brain tissue samples from 1,719 autopsy cases were compared.

Metabolomics-driven determination of targets for salicylic acid and ibuprofen in positive electrospray ionization using LC-HRMS.

Due to the large number of basic therapeutic and illicit drugs, systematic toxicological analysis has widely been performed with liquid chromatography coupled to mass spectrometry using positive electrospray ionization. However, there exist a smaller number of drugs, typically acidic drugs, which require the use of negative electrospray ionization either via a separate analysis or polarity switching. Here, targets relating to salicylic acid and ibuprofen in positive electrospray ionization were determined through a metabolomics-driven retrospective investigation of forensic casework.

Development of a single retention time prediction model integrating multiple liquid chromatography systems: Application to new psychoactive substances.

Database-driven suspect screening has proven to be a useful tool to detect new psychoactive substances (NPS) outside the scope of targeted screening; however, the lack of retention times specific to a liquid chromatography (LC) system can result in a large number of false positives. A singular stream-lined, quantitative structure-retention relationship (QSRR)-based retention time prediction model integrating multiple LC systems with different elution conditions is presented using retention time data (n = 1281) from the online crowd-sourced database, HighResNPS. Modelling was performed using an artificial neural network (ANN), specifically a multi-layer perceptron (MLP), using four molecular descriptors and one-hot encoding of categorical labels.

Cocaine profiling method retrospectively developed with nontargeted discovery of markers using liquid chromatography with time-of-flight mass spectrometry data.

Illicit drug profiling performed by forensic laboratories assists law enforcement agencies through providing information about chemical and/or physical characteristics of seized specimens. In this article, a model was developed for the comparison of seized cocaine based on retrospective analysis of data generated from ultrahigh performance liquid chromatography with time-of-flight mass spectrometry (UHPLC-TOF-MS) comprehensive drug screening. A nontargeted approach to discover target compounds was employed, which generated 53 potential markers using data from cocaine positive samples.

Simple implementation of muscle tissue into routine workflow of blood analysis in forensic cases - A validated method for quantification of 29 drugs in postmortem blood and muscle samples by UHPLC-MS/MS.

Whole blood is most often the matrix of choice for postmortem analysis but it is not always available. In these cases, muscle tissue can be used as an alternative matrix. Therefore, an ultra-high-performance liquid chromatography-tandem mass spectrometry method for the quantification of 29 drugs and metabolites of toxicological interest in postmortem muscle tissue was developed and validated.

Analysis of seized peptide and protein-based doping agents using four complimentary methods: Liquid chromatography coupled with time of flight mass spectrometry, liquid chromatography-ultraviolet, Bradford, and immunoassays.

Analysis and identification of seized doping-related products are important tasks for customs or forensic laboratories in order to prevent potentially dangerous and illegal compounds to go into circulation. At the Section of Forensic Chemistry in Copenhagen, we have a workflow consisting of four complimentary validated methods to identify common doping-related substances: liquid chromatography-ultraviolet (LC-UV), LC coupled with time of flight mass spectrometry (LC-TOF-MS), the colorimetric Bradford assay, and an immunoassay. The Bradford assay screens for peptide or proteins in the sample, and the immunoassay confirmed human chorionic gonadotropin (hCG).

Identification of phenobarbital and other barbiturates in forensic drug screening using positive electrospray ionization liquid chromatography-high resolution mass spectrometry.

Comprehensive drug-screening performed by liquid chromatography-high resolution mass spectrometry (LC-HRMS) enables identification of hundreds to thousands of drug compounds in a single analysis. Forensic drug screening is generally performed with positive electrospray ionization (ESI ), targeting basic drugs; however, a few toxicologically important drugs such as barbiturates, may require analysis by negative ESI. In this work, screening targets for barbiturates were determined using our LC-HRMS screening with ESI .

Retrospective analysis for valproate screening targets with liquid chromatography-high resolution mass spectrometry with positive electrospray ionization: An omics-based approach.

Liquid chromatography coupled with high-resolution mass spectrometry (LC-HRMS) is an important analytical tool in the systematic toxicological analysis performed in forensic toxicology. However, some important compounds, such as the antiepileptic drug valproate (valproic acid; VPA), cannot be directly detected with positive electrospray ionization (ESI ) due to poor ionization. Here we demonstrate an omics-based retrospective analysis for the identification of indirect screening targets for VPA in whole blood with LC-ESI -HRMS.

Glycine-modified growth hormone secretagogues identified in seized doping material.

A number of unknown pharmaceutical preparations seized by Danish customs authorities were submitted for liquid chromatography-high resolution mass spectrometry (LC-HRMS) analysis. Comparison with reference standards unequivocally identified the content of the powders as analogs of the growth hormone secretagogues GHRP-2 (Pralmorelin), GHRP-6, Ipamorelin, and modified growth hormone releasing factor (modified GRF 1-29), which can be used as performance-enhancing substances in sports. In all cases, the detected modification involved the addition of an extra glycine amino acid at the N-terminus, and analytical methods targeting growth hormone secretagogues should hence be updated accordingly.

Driving under the influence of alcohol and drugs in the eastern part of Denmark in 2015 and 2016: Abuse patterns and trends.

Objectives: The objective of this study was to examine the frequency of psychoactive drugs and alcohol in drivers under suspicion of driving under the influence of drugs and alcohol in 2015 and 2016 in the eastern part of Denmark. The trends in the number of traffic cases sent for drug analysis since 2000 and alcohol analysis since 2011 are also discussed.

Methods: Blood samples from drivers suspected of being under the influence of alcohol and/or medication and/or illicit drugs in 2015 and 2016 were investigated as requested by the police.

Postmortem concentrations of gamma-hydroxybutyrate (GHB) in peripheral blood and brain tissue - Differentiating between postmortem formation and antemortem intake.

Gamma-hydroxybutyrate (GHB) is a recreational drug, a drug of abuse, as well as an endogenous molecule in mammals. The drug has become infamous as a tool for drug-facilitated sexual assault. GHB is found in low concentrations in living humans, while at postmortem the concentration of GHB rises due to fermentation processes.

Screening for illicit and medicinal drugs in whole blood using fully automated SPE and ultra-high-performance liquid chromatography with TOF-MS with data-independent acquisition.

A broad forensic screening method for 256 analytes in whole blood based on a fully automated SPE robotic extraction and ultra-high-performance liquid chromatography (UHPLC) with TOF-MS with data-independent acquisition has been developed. The limit of identification was evaluated for all 256 compounds and 95 of these compounds were validated with regard to matrix effects, extraction recovery, and process efficiency. The limit of identification ranged from 0.

Quantification of 31 illicit and medicinal drugs and metabolites in whole blood by fully automated solid-phase extraction and ultra-performance liquid chromatography-tandem mass spectrometry.

An efficient method for analyzing illegal and medicinal drugs in whole blood using fully automated sample preparation and short ultra-high-performance liquid chromatography-tandem mass spectrometry (MS/MS) run time is presented. A selection of 31 drugs, including amphetamines, cocaine, opioids, and benzodiazepines, was used. In order to increase the efficiency of routine analysis, a robotic system based on automated liquid handling and capable of handling all unit operation for sample preparation was built on a Freedom Evo 200 platform with several add-ons from Tecan and third-party vendors.