Interpretation of postmortem toxicology results is complicated by postmortem redistribution (PMR), which can alter drug concentrations between tissues and blood (BL). This study evaluates brain (BR) to BL concentration ratios for amphetamine (AMP), methamphetamine (MAMP), cocaine (COC), benzoylecgonine (BE), cocaethylene (CE), and fentanyl (FENT) using postmortem casework data to evaluate the utility of BR tissue as an alternative matrix. BR:BL ratios varied widely across analytes: AMP (2.2-5.8), BE (0.2-2.1), CE (1.2-5.1), COC (0.1-5.8), MAMP (2.2-7.3), and FENT (1.1-24.8). Lipophilic compounds (AMP, CE, COC, MAMP, FENT) generally exhibited ratios greater than one, consistent with preferential BR partitioning and susceptibility to PMR, whereas the polar metabolite BE showed ratios less than one, reflecting limited BR accumulation. FENT demonstrated the greatest variability, highlighting challenges in interpreting its postmortem concentrations. Correlations between BR and femoral BL concentrations were slightly positive for all analytes, indicating that BR concentrations may not be reliable proxies for BL levels. Evaluation of select antemortem cases and outliers revealed additional influences on BR:BL ratios, including survival time, drug stability, route of administration, and polysubstance use. COC instability and hydrolysis to BE, as well as incomplete drug distribution in rapid deaths, contributed to atypical ratios. Overall, BR:BL ratios are highly analyte dependent and influenced by both pharmacokinetic properties and postmortem factors. While BR tissue may provide useful complementary information when BL is unavailable, it should not be used as a direct substitute for BL in toxicological interpretation. These findings underscore the need for further research to expand BR:BL datasets across drug classes.

Drug-facilitated sexual assault (DFSA) refers to sexual acts where the victim is incapacitated by the effects of psychotropic substances, causing impaired ability to prevent an attack and/ or provide informed consent. To date, the level of understanding of the prevalence of the specific drugs involved in DFSA has been limited. This review examines global toxicological case data to assess the prevalence of tramadol, a widely prescribed opioid analgesic, in the context of DFSA. Tramadol was confirmed in reports across Africa, Oceania, Europe, the Middle East, and North America, with detection rates ranging from 1-33%. In these cases, 17 voluntary and involuntary drug combinations were identified including alcohol, opioids, benzodiazepines, Z-drugs, cannabinoids and stimulants. These substances pose significant risk for both pharmacodynamic and pharmacokinetic interaction with tramadol, potentially altering plasma metabolite levels, clearance times and also central nervous system depression, facilitating DFSA and influencing forensic detection and interpretation of tramadol and its metabolites. Overall, this review underscores the significance of tramadol in DFSA casework and highlights the importance of drug-drug interactions in order to improve forensic analysis in DFSA casework and aid in legal action leading to victim support and closure.

Treatment with antipsychotics such as chlorprothixene, clozapine, olanzapine and quetiapine can be associated with cardiovascular and metabolic risks and poisoning, making them highly relevant in toxicological investigations. Since antipsychotics primarily act within the central nervous system, brain tissue serves as an alternative or complementary matrix to femoral blood in toxicological analysis. This study aimed to retrospectively evaluate brain concentrations and brain-blood ratios across toxic and non-toxic levels in 466 postmortem cases involving chlorprothixene, clozapine, olanzapine and/or quetiapine. Median concentrations in brain tissue across all cases were 0.20 mg/kg for chlorprothixene, 4.2 mg/kg for clozapine, 0.42 mg/kg for olanzapine and 0.34 mg/kg for quetiapine. Corresponding brain-blood ratios remained stable across the examined concentration range with median ratios of 3.1 for chlorprothixene, 2.9 for clozapine, 2.6 for olanzapine and 3.0 for quetiapine. The provided brain concentrations and ratios can support further toxicological evaluations of postmortem cases.

We report a case of a previously healthy 3-year-old girl who developed acute altered consciousness, agitation, and visual hallucinations consistent with an anticholinergic toxidrome. Initial examinations were unremarkable, and urine immunoassay screening (SIGNIFY™ ER) was negative. Serum analysis by liquid chromatography-quadrupole time-of-flight mass spectrometry (LC-QTOF/MS) identified chlorpheniramine and scopolamine, while excluding other agents. Concentrations were 370 ng/mL for chlorpheniramine and 1 ng/mL for scopolamine, both near reported toxic ranges. An empty blister pack of 22 pediatric motion sickness tablets was found at the patient's home, confirming accidental overdose of an over-the-counter (OTC) anti-motion sickness medication. This case demonstrates the diagnostic utility of high-resolution mass spectrometry in pediatric poisoning, particularly for drugs not detected by routine screening. It represents a rare report documenting quantitative confirmation of both chlorpheniramine and scopolamine in a child, with correlation between serum concentrations and clinical presentation. Scopolamine is highly toxic even at low concentrations, and pediatric cases with measured blood levels are exceedingly rare. Our findings highlight the need to consider additive or synergistic toxicity in OTC formulations containing multiple anticholinergic agents and emphasize the importance of caregiver education regarding the potential risks of OTC medications often perceived as safe.

Ceramides and sphingosine-1-phosphate (S1P) are bioactive sphingolipids critically involved in the regulation of cell survival and death, whose homeostasis is disrupted by alcohol exposure. A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for the simultaneous quantification of nine ceramide species and S1P in serum. Following protein precipitation, analytes were separated on a CSH C18 column within an 11-min chromatographic run. Calibration curves were linear over 5 to 500 ng/mL with R2 greater than 0.99 for all analytes. Intra- and inter-day accuracy and precision met acceptance criteria across all quality control levels. Extraction recoveries ranged from 79.51% to 109.23%, matrix effects were adequately compensated by stable isotope-labeled internal standards, and dilution integrity was confirmed across factors of 1:5, 1:10, and 1:20. The method was applied to a large cohort of serum samples from suspected alcohol-impaired driving cases stratified by blood alcohol concentration (BAC) into negative control, mild, moderate, and severe intoxication groups. Very long-chain ceramides including Cer 22:0, Cer 24:0, and Cer 24:1, as well as S1P, were consistently elevated in alcohol-exposed groups relative to the negative control. These findings indicate that circulating sphingolipid profiles reflect lipid metabolism alterations induced by ethanol exposure and may provide complementary information to blood alcohol concentration in the assessment of alcohol-associated cases.

Acute recreational drug toxicity (ARDT) is a frequent cause of emergency department (ED) presentations, yet comprehensive toxicological testing by chromatographic techniques is rarely conducted due to limited availability and resource-intensive workflows. Consequently, initial clinical decisions are made without analytical confirmation. Paper spray-mass spectrometry (PS-MS) may offer an alternative approach for targeted clinical toxicology screening owing to its rapid and simplified analysis of biological samples. Serum donor samples from patients presenting to the ED with suspected ARDT were analyzed using PS-MS and liquid chromatography-tandem mass spectrometry (LC-MS/MS) for method comparison. Targeted screening and quantitative performance were evaluated using pooled diagnostic metrics, orthogonal regression, per-analyte % bias and Bland-Altman analyses. PS-MS was subsequently applied to a larger screening-only cohort to assess analytical performance, detection frequency, and concentration distributions across a clinically relevant drug panel. Across 114 research samples, PS-MS demonstrated high targeted screening performance, with a pooled sensitivity of 82.7%, specificity and positive predictive value of 100%, and negative predictive value of 98.2%. Quantitative comparison showed strong correlation between methods, with proportional bias at higher concentrations and no single analyte influencing the agreement metrics. In the screening-only cohort (n = 99), PS-MS detected and quantified a broad range of analytes, including evidence of polydrug use. Several low-frequency analytes were not observed within the analyzed serum donor sample cohort. PS-MS may provide a fast and specific targeted screening approach for ARDT in ED samples, delivering analytically informative toxicological data. While not a replacement for confirmatory testing or a validated in vitro diagnostic assay, this research study into PS-MS represents a valuable complementary tool to support downstream confirmatory testing.

Post-mortem forensic toxicology aims to assist in cause-of-death investigations, especially in suspected toxicity cases. Utilizing a large toxicology database derived from routine medico-legal autopsies, we tested a probabilistic approach in assessing the contribution of a toxicological finding to the underlying cause of death. In this retrospective study, theoretical general prior odds for a poisoning death were calculated for three different groups of cases based on the laboratory referral issued by the forensic pathologist. The groups were, in descending order according to the likelihood of poisoning: (i) suspected poisoning (N = 6356), (ii) exclusion of poisoning (N = 12 641), and (iii) for background information only (N = 15 018). Propranolol was selected as a model compound with 548 propranolol-positive cases in the database. First, the detected propranolol concentrations were divided into concentration ranges. Then, a likelihood ratio (LR) for each concentration range was calculated as the probability of observing a propranolol concentration within a certain concentration range given that propranolol poisoning was the cause of death, divided by the probability of observing a propranolol concentration within the same concentration range given that propranolol poisoning was not the cause of death. To showcase the practical applicability, posterior odds for fatal propranolol poisoning were derived using the odds form of Bayes' theorem (posterior odds = prior odds × LR) for a selection of propranolol-positive cases (N = 5) and compared with the cause of death recorded on the death certificate. In our dataset, the point estimate for LR exceeded 1 for propranolol concentrations ≥1.0 mg/L. The LR proved useful in assisting in the cause-of-death determination and making the interpretive process more systematic and transparent. Despite some limitations, this study added to the evidence of the usefulness of a probabilistic approach in assessing the relevance of a toxicology finding in autopsy cases.

Drug-facilitating sexual assault (DFSA) corresponds to sexual activity occurring whereas the victim is unable to express consent due to the action of drugs. Although the use of sedative and amnesic drug in DFSA, such as benzodiazepines and gamma-hydroxybutyrate, is widely described in the medical literature, there is a lack of information regarding the involvement of amphetamine derivatives in this context, particularly synthetic cathinones. Herein, we report a case of a 19-year-old woman who presents herself to the emergency department following an acute alcoholic intoxication and a suspected DFSA after going to a nightclub. The emergency toxicological screening on the plasma and urine of the patient, performed through liquid chromatography coupled to high resolution tandem mass spectrometry in target and non-target mode and with gas chromatography coupled to high resolution mass spectrometry, enabled the detection of 4-bromomethcathinone, a new psychoactive substance (NPS) of the cathinone family. The demethylated and reduced metabolites of the drug were also found in both plasma and urine. No other substance was identified except ethanol, which was quantified in gas chromatography-flame ionization detection, and diazepam with its metabolites, this last one having been administrated in the emergency department. As far as we know, this report described the first case of DFSA involving this halogenated derivative of cathinone and consequently, highlights the need for toxicologists to systematically investigate with specific analytical methods the potential presence of NPS in cases of DFSA.

The structural similarity of positional isomers of fentanyl analogs makes their forensic differentiation difficult, particularly when the parent compounds exhibit similar chromatographic behavior and product-ion spectra. This study evaluated the differentiation of 2-, 3-, and 4-fluorofuranylfentanyl (2-, 3-, and 4-FFF) using high-performance liquid chromatography-mass spectrometry (HPLC-MS) and human liver microsomes (HLMs). The synthesized positional isomers were analyzed using gas chromatography-mass spectrometry (GC-MS), high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS), and HPLC-ion trap-time-of-flight mass spectrometry (HPLC-IT-TOF-MS). Although the parent isomers were separated by GC-MS, their retention times and product-ion spectra obtained by HPLC-MS/MS were highly similar under the present conditions. Incubation with HLMs supplemented with NADPH, UDPGA, and alamethicin produced three Phase I metabolic categories: an oxidative N-dealkylated metabolite of FFF (M1), monohydroxylated FFF metabolites (M2a and M2b), and an amide hydrolysis product of FFF (M3). Glucuronide conjugates were not detected under the present analytical conditions. Although M1 was the predominant metabolite, M1 did not retain the complete FFF structure. M2a and M2b were putatively assigned as monohydroxylated metabolites that retained the complete FFF structure, and may therefore support the differentiation of FFF positional isomers, although their retention times did not allow the differentiation. M3, which did not retain the complete FFF structure, showed distinct retention times among the three positional isomers, providing chromatographic information useful for metabolite-based differentiation. The in vitro elimination half-lives differed among 2-, 3-, and 4-FFF under the present HLM conditions, suggesting possible differences in the pharmacokinetic properties of these positional isomers. These findings indicate that HPLC-MS analysis using HLMs provides useful analytical information for differentiating FFF positional isomers.

This study describes the contextual factors and scientific data associated with fentanyl-related deaths among individuals aged 0-17 years in North Carolina. Using information from the North Carolina Medical Examiner System from 2015 to 2024, 138 pediatric fentanyl deaths were identified. Manually abstracted scene information, toxicology results, and pathological findings allowed for a rarely examined in-depth analysis of circumstances surrounding these deaths. Children less than 2 years (n = 33, 1.38 per 100 000) and adolescents aged 15-17 years (n = 80, 1.96 per 100 000) experienced a fentanyl-related death rate higher than the statewide pediatric fentanyl-related death rate of 0.60 per 100 000 residents. Among children less than 2 years, 78.8% of overdose onset occurred in the decedent's primary residence, and drugs or paraphernalia were present at the scene in 51.5% of cases. Co-sleeping was documented in 60.6% of deaths, with 80.0% occurring in adult beds. Postmortem blood concentrations of fentanyl among children less than two years ranged from less than 0.50 to 280 ng/mL (mean 27.0 ng/mL, median 17 ng/mL). Among adolescents, 62.5% of overdose onset occurred in the decedent's primary residence, followed by a friend's residence (13.8%). History of substance use was documented in 82.5% of cases and drugs or paraphernalia were present in 67.5% of cases. Blood concentrations of fentanyl among adolescents ranged from less than 1.0 to 330 ng/mL (mean 21.7 ng/mL, median 11.0 ng/mL). Scene investigation and documentation allow for the recognition of behavioral trends, which can be targeted for age-specific overdose prevention strategies. Among decedents less than 2 years, notable trends included children co-sleeping in adult beds with access to drugs and drug waste and/or prolonged periods without supervision. Among older adolescents, several cases involved suspected counterfeit pharmaceuticals and/or periods of non-contact with primary caregivers. Improved workflows for pediatric medicolegal death investigation and electronic reporting strategies are already underway.

Motor vehicle fatalities continue to be a leading cause of preventable death in the United States. Beginning in 2019, there was a steady uptick in traffic fatalities nationally, which peaked in 2021. The state of Connecticut has experienced similar trends with increases in traffic fatalities over the last several years. The focus of this review was to evaluate drug positivity in traffic-related fatalities that occurred in Connecticut between January 1, 2020 and December 31, 2024. Cases related to traffic fatalities were identified by the state's Office of the Chief Medical Examiner. Deaths of drivers, passengers, and pedestrians were used for the data set. A total of 1,816 fatal motor vehicle incidents were reviewed. Of these, 312 cases lacked toxicology results, and 5 cases had testing canceled; both were excluded from substance-related analyses. Among the remaining 1,499 cases with available toxicological data, 1,084 (72%) had at least one positive toxicology finding. Drug positivity was identified in 75% (710 of 948) of driver fatalities, 66% (125 of 188) of passenger fatalities, and 67% (221 of 327) of pedestrian fatalities. The most frequently encountered drugs across all three classes of decedents were ethanol, delta-9 tetrahydrocannabinol, and fentanyl. These findings underscore the pervasive role of drugs and alcohol across all categories of traffic-related deaths and highlight the ongoing need for preventive strategies, toxicology testing, and targeted public safety interventions.

Interpretation of postmortem (PM) γ-hydroxybutyric acid (GHB) remains challenging due to endogenous production and PM increases in concentration associated with decomposition. Existing interpretation guidelines primarily rely on a single criterion, such as concentration thresholds or urine-to-blood ratios. In San Francisco, routine and unbiased implementation of GHB testing across all casework generated a large dataset including 1865 GHB detections in PM blood or urine, of which 552 were paired peripheral blood and urine samples. Cases were categorized as acute exogenous use, confirmed exogenous use, or no indication of use based on toxicological and investigative findings. Peripheral blood concentrations in cases with no indication of use did not vary significantly by age, gender, race, or manner of death but were significantly higher in decomposed cases. While existing guidelines effectively identify endogenous levels, the proposed hybrid model, combining peripheral blood concentration (B) with urine-to-blood ratio (U/B), enables further classification into no exogenous use, possible exogenous use (B > 5 mg/L and U/B > -0.05 × B + 4), and acute exogenous use (B > 50 mg/L and U/B > -0.05 × B + 10). The model was evaluated and compared to existing guidelines. Internal applicability was assessed using recent casework, and external applicability was evaluated using an independent dataset from Québec, Canada. The hybrid model demonstrated improved performance, achieving substantially higher positive predictive value when compared to existing guidelines, while maintaining equal or improved performance measures across other metrics. This approach provides a conservative, evidence-based framework for distinguishing acute exogenous GHB use in PM casework.

Drug-facilitated sexual assault (DFSA) is a type of drug-facilitated crime defined as a form of sexual violence against an individual incapacitated or unconscious due to the effects of psychoactive substances. These substances alter the victim's level of consciousness, judgment, and/or memory. This study aimed to develop and validate a quantitative LC-MS/MS method for psychoactive substances and biotransformation products with detection limits suitable for DFSA investigations. The scope was based on ANSI/ASB Standard 121 guidelines (2021). Urine samples were submitted to enzymatic hydrolysis, followed by liquid-liquid extraction. An aliquot of urine underwent protein precipitation with acetonitrile. After agitation, centrifugation, and evaporation of solvent, samples were resuspended with mobile phase and injected into LC-MS/MS system. The method was validated according to ANSI/ASB Standard 036 recommendations, with quantification limits of 0.5 to 50 ng/mL and linearity of 0.5 to 750 ng/mL. Bias and imprecision were better than 14.3% and 14.8%, respectively, and matrix effect ranging from -72.9% to 21.5%. No evidence of carryover and interference was observed. All substances were stable at 10 °C for 24 hours and recovery results were better than 5.2%. In total, 42 samples from sexual assault victims were analyzed, with at least one substance detected in 26 samples. Ethanol was the most predominant substance, followed by 11-nor-9-carboxy-Δ-9-tetrahydrocannabinol and cocaine. A sensitive method was developed and validated to quantify psychoactive substances in urine, with low limits of detection and quantification, adequate linearity, bias, imprecision, and successfully applied to the analysis of authentic urine samples.

The performance of the SAMElife™ rapid hair screening kit was evaluated for the detection of drugs of abuse in 31 anonymized hair samples, encompassing 217 analyses for morphine, cocaine, amphetamines, buprenorphine, oxycodone, fentanyl, and methadone. Screening results were compared with confirmatory gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-tandem mass spectrometry (LC-MS/MS) analyses. The kit demonstrated high specificity (100%) and good sensitivity (81.8%) for basic drugs, with four false negatives observed for cocaine, while no false positives were recorded. Detection of Δ9-tetrahydrocannabinol (THC) was unreliable, yielding uniformly positive results regardless of confirmatory analysis. The automated workflow, requiring only hair cutting and no prior decontamination, provides a rapid and practical approach for preliminary screening in routine toxicology laboratories. Despite limitations in THC detection and the small number of positive samples for some analytes, the SAMElife™ kit offers a reliable tool to reduce the number of samples requiring confirmatory analysis. Further evaluation on larger populations and optimization of the THC assay are recommended.