Journal Of Analytical Toxicology[JOURNAL]
Sun
200 papers
Determination of specific absorbance (A¦) for four designer benzodiazepines.
Ku BS, Vargas JR
J Anal Toxicol · 2026 May · PMID 41863317 · Full text
Specific absorbance (A¦) is a unitless value defined as the absorbance of a 1% (w/v) solution measured at a 1 cm path length. Establishing reliable specific absorbance values allows forensic toxicology laboratories to ve...
Specific absorbance (A¦) is a unitless value defined as the absorbance of a 1% (w/v) solution measured at a 1 cm path length. Establishing reliable specific absorbance values allows forensic toxicology laboratories to verify the concentration of analytical stock solutions. Designer benzodiazepines have appeared with increasing frequency in forensic casework but often lack validated reference values. This study determined specific absorbance values for bromazolam, etizolam, flualprazolam, and flubromazepam using ultraviolet-visible spectroscopy in an aqueous acidic solution. Measurements were collected in duplicate at three concentrations across three days at the wavelength of maximum absorbance for each compound. Inter-run and intra-run variability were evaluated using the coefficient of variation, and the reliability of the values was classified according to Clarke's Analysis of Drugs and Poisons criteria. The experimentally derived values reported here provide reference information that may assist laboratories in verifying analytical standards for emerging benzodiazepines.
Case-control study of mitragynine deaths in Florida.
Iuteri CM, Sawyers RJ, Byrne CM … +2 more , Trojano M, Thundiyil J
J Anal Toxicol · 2026 May · PMID 41837384 · Publisher ↗
Kratom use has increased in the USA, with its primary alkaloid, mitragynine, detected in postmortem toxicology, and implicated in reported fatalities. This study evaluated toxicologic concentrations and autopsy findings...
Kratom use has increased in the USA, with its primary alkaloid, mitragynine, detected in postmortem toxicology, and implicated in reported fatalities. This study evaluated toxicologic concentrations and autopsy findings associated with deaths attributed to mitragynine. Autopsy and postmortem toxicology reports were obtained from the majority of Florida medical examiner districts over 5 years for decedents in whom mitragynine was listed as a cause of death (mitragynine-induced fatalities). A comparison group included cases from a single county in which mitragynine was detected but not deemed causal (mitragynine-associated deaths). Demographic, toxicologic, and autopsy data were abstracted using standardized instruments. Thirty-eight mitragynine-induced fatalities and 111 mitragynine-associated deaths were identified, with similar demographic characteristics between groups. Mitragynine-associated deaths all featured co-exposures, including fentanyl (n = 84), cocaine (n = 49), and ethanol (n = 29). Mitragynine-induced fatalities had no toxic levels of any other substance and demonstrated higher rates of pulmonary edema (55.6% vs 31.5%), hepatomegaly (47.4% vs 14.4%), minor facial trauma (18.4% vs 5.4%), and cardiomegaly (47.4% vs 11.8%). Mean blood mitragynine concentrations were substantially higher in mitragynine-induced fatalities (2486 µg/L; 95% CI 1607-2863) compared with mitragynine-associated deaths (179 µg/L; 95% CI 217-375), suggesting a concentration-dependent association with fatal toxicity. Mitragynine toxicity accounts for a notable number of deaths.
HPLC-MS/MS stereoselective determination and quantification of MDMA and its phase-1 metabolite in human oral fluid samples: estimation of consumption time.
Balloni A, Wille SMR, Sprega G … +3 more , Kobidze G, Tini A, Lo Faro AF
J Anal Toxicol · 2026 Mar · PMID 41837368 · Publisher ↗
3,4-Methylenedioxymethamphetamine (MDMA), commonly known as Ecstasy, is a psychoactive substance used for recreational purposes. This study used a stereoselective method based on high-performance liquid chromatography co...
3,4-Methylenedioxymethamphetamine (MDMA), commonly known as Ecstasy, is a psychoactive substance used for recreational purposes. This study used a stereoselective method based on high-performance liquid chromatography coupled with triple quadrupole mass spectrometry (HPLC-MS/MS) to quantify MDMA and 3,4-methylenedioxyamphetamine (MDA) enantiomers in human oral fluid (OF). Samples were collected during a clinical trial and roadside police checks. The enantiomer concentrations were determined in 161 OF samples. Pharmacokinetic data showed that the S-(+)-enantiomer of MDMA was eliminated faster than the R-(-)-enantiomer, with half-lives of 3.3 hours and 4.8 hours, respectively. Peak concentrations for both enantiomers were reached within 1.75 hours post-administration. For MDA, the 2nd-enantiomer was eliminated more quickly, with a half-life of 9.6 hours compared to 23.8 hours for the 1st-enantiomer. The analysis of the MDMA R/S enantiomer ratio revealed a gradual increase over time, suggesting the possibility of estimating the time since ingestion. In samples collected during roadside checks, 54.5% of cases had an R/S ratio greater than 1.5, which might indicate consumption within the previous 24 hours. In conclusion, OF proved to be a promising non-invasive matrix for the detection of MDMA and MDA. The stereoselective analysis provided useful information for estimating the time of substance intake, especially in forensic contexts such as Driving Under the Influence of Drugs (DUID) cases.
Psilocin glucuronide in whole blood: a stable and useful biomarker of psilocybin intake.
Bergh MS, Bogen IL, Vevelstad M … +1 more , Øiestad ÅML
J Anal Toxicol · 2026 May · PMID 41723823 · Full text
Detecting psilocybin use is challenging because it rapidly converts to its psychoactive metabolite psilocin, and both compounds are unstable in blood. Bufotenin, a structural isomer of psilocin, may exhibit comparable in...
Detecting psilocybin use is challenging because it rapidly converts to its psychoactive metabolite psilocin, and both compounds are unstable in blood. Bufotenin, a structural isomer of psilocin, may exhibit comparable instability in blood. For reliable detection, we developed and validated an LC-MS/MS method to simultaneously quantify psilocin, bufotenin, and their metabolites, psilocin glucuronide (PSG) and 5-hydroxyindole-3-acetic acid (5-HIAA), in human whole blood. We prepared blood samples by protein precipitation and lipid removal filtration. Analytes were separated using a biphenyl column. The method was validated according to AAFS guidelines with LOQs of 2.4 nM for psilocin, PSG, and bufotenin, and 30 nM for 5-HIAA. We assessed analyte stability in whole blood under conditions relevant to forensic sample handling. Psilocin degraded by 46%-66% at room temperature and 66%-76% at 4°C after one day, increasing to 88%-99% and 94%-100% after three days. At -20°C, degradation slowed, with up to 51% loss after one month and ≥91% after three months. In contrast, PSG remained stable for 14 days at both room temperature and 4°C, and for at least one year at -20°C, making it a reliable biomarker of psilocybin intake. Bufotenin showed moderate stability, while 5-HIAA was unsuitable as a biomarker due to its endogenous presence. Our method enables direct quantification of PSG, offering a straightforward and accurate alternative to indirect approaches. We demonstrated the method's applicability by analyzing 23 forensic blood samples that screened positive for psilocin or PSG, with PSG quantified in nearly all cases, even when psilocin was below LOQ. These findings confirm PSG as a specific and stable biomarker of psilocybin use, and its integration into routine forensic workflows could significantly improve detection reliability.
Simultaneous analysis of Δ9-THC, Δ8-THC, CBD, and CBN in breath aerosols collected using cannabix technologies breath collection unit.
Deeb S, Fabian ZE, Määttä M … +2 more , Fraccarolli P, Engelhart DA
J Anal Toxicol · 2026 May · PMID 41723813 · Full text
A simple, accurate, and cost-effective liquid chromatography-tandem mass spectrometric (LC-MS/MS) method has been developed and validated for the simultaneous quantification of Δ9-tetrahydrocannabinol (Δ9-THC), Δ8-tetrah...
A simple, accurate, and cost-effective liquid chromatography-tandem mass spectrometric (LC-MS/MS) method has been developed and validated for the simultaneous quantification of Δ9-tetrahydrocannabinol (Δ9-THC), Δ8-tetrahydrocannabinol (Δ8-THC), cannabidiol (CBD), and cannabinol (CBN) in breath aerosols collected using the Cannabix Technologies' Breath Collection Unit (BCU). The method is suitable for use in workplace drug testing and in the investigation of driving under the influence of drugs (DUID) cases. The method was validated according to ANSI/ASB Standard 036, First Edition 2019 guidelines. A lower limit of quantification (LLOQ) of 2.5 pg/L was achieved. Calibration curves were linear (R2 > 0.995) and quality control (QC) accuracy between 91.3% and 100.5% with precision (%CV) less than 17.7%. No major matrix effects or drug interferences were observed. Samples were stable at room temperature and refrigerated at 2-8°C up to 7 days. Breath samples were collected using the BCU from volunteer regular cannabis users before cannabis consumption and at multiple timepoints after smoking cannabis up to 90 minutes. The results verified the methods ability to capture and quantitate the cannabinoids in breath. The data indicated similar concentration versus time trends for THC metabolism assessed from breath to that previously reported from blood. Breath is an alternative, non-invasive sample matrix that holds promise for identifying recent cannabis use.
Unlocking the potential of bone marrow in postmortem forensic toxicology: current evidence and future perspectives.
Vanerio S, Galante N, Ravelli A … +3 more , Bergamaschi RF, Battistini A, Casati S
J Anal Toxicol · 2026 Jun · PMID 41719197 · Full text
Bone marrow (BM) has emerged as a valuable alternative matrix in postmortem toxicology, when conventional samples such as blood or soft tissues are unavailable, degraded or contaminated. BM, due to its protecting anatomi...
Bone marrow (BM) has emerged as a valuable alternative matrix in postmortem toxicology, when conventional samples such as blood or soft tissues are unavailable, degraded or contaminated. BM, due to its protecting anatomical site within the medullary cavity, slows down decomposition, limits environmental and microbial interference and enables the preservation of xenobiotics over extended postmortem intervals. This review summarizes the updated literature on BM anatomy, sampling site, xenobiotic distribution and stability, as well as the influence of postmortem redistribution (PMR) and putrefaction for toxicological interpretation of the analytical results. Evidence indicates that drug distribution in BM is governed by tissue vascularity and analyte physicochemical properties, with lipophilic compounds often reaching higher concentrations in BM than in blood. Numerous studies demonstrate the long-term stability of drugs such as amphetamines, benzodiazepines, sedative-hypnotics and ethanol in BM, even in severely decomposed, burned, or skeletonized remains. While BM shows reduced susceptibility to PMR and putrefaction compared to blood and soft tissues, post-collection stability depends on proper storage conditions. Findings from human forensic cases assess the detectability of a wide range of illicit and therapeutic substances in BM, supporting its utility in reconstructing ante-mortem drug exposure when traditional matrices are compromised. Despite promising results, data remain limited, and further research is needed to refine interpretative frameworks and establish standardized protocols for BM sampling, analysis, and toxicological evaluation.
Illicit opioid adulterant trends in patients presenting with acute opioid overdose.
Levine M, Culbreth R, Buchanan J … +18 more , Schwarz E, Love J, Brent J, Meaden CW, Judge B, Aldy K, Hendrickson R, Hughes A, Krotulski A, Logan B, Shulman J, Carpenter JE, Wax P, Maducci AM, Calello DP, Campleman S, Manini AF, Toxicology Investigators Consortium (ToxIC) Fentalog Study Group
J Anal Toxicol · 2026 May · PMID 41712495 · Publisher ↗
Clandestine fentanyl manufacturing oftentimes introduces adulterants and contaminants. This article aims to evaluate trends in adulterants from a cohort of patients presenting to the emergency department (ED) with illici...
Clandestine fentanyl manufacturing oftentimes introduces adulterants and contaminants. This article aims to evaluate trends in adulterants from a cohort of patients presenting to the emergency department (ED) with illicit opioid overdose across the United States. The Fentalog Study group is a multicenter toxicology study group which evaluated ED patients with suspected opioid overdose at 10 medical centers across the United States between 21 September 2020 through 5 February 2024. Comprehensive qualitative toxicology testing was performed on residual serum specimens. Study sites were divided into three geographic regions: West (California, Oregon, Colorado), Midwest (Missouri, Michigan), and East (New York, New Jersey, Pennsylvania, Georgia). Illicit opioids were defined as fentanyl and fentanyl analogs, heroin or its metabolites, and/or novel potent opioids such as nitazenes. 1295 patients with confirmed illicit opioid overdose were included. Males accounted for the majority (73.7%) of patients. The median age was 39 (IQR: 31-54) years. Adulterants were detected in 745 (57.5%) patients. Quinine was the most abundantly encountered adulterant (433; 33.4%). Antihistamines were the most frequently detected class of adulterants (19.6%). There were significant differences in adulterants detected across the three time periods, with notable decreases in adulterants from time-period 1 (79.5%) to time-period 3 (41.7%; P < .001). Adulterants were found in 84 (27.0%) of patients that presented to a hospital in the Western United States, compared with 181 (24.3%) in the Midwest, and 480 (64.4%) of patients in the East (P < .001). Patients with concurrent cocaine were more likely having an adulterant present than those without cocaine present (OR 1.23; 95% CI 1.15-1.31). In contrast, patients with illicit opioids and concurrent methamphetamine were less likely to have adulterants present (OR 0.89; 95% CI 0.84-0.95). Adulteration of illicit opioids was more likely in the Eastern United States and for those with concurrent cocaine and opioid exposures.
GC-FID quantification of methanol, ethanol, 2-propanol, acetone, ethylene glycol, diethylene glycol, 1,2-propylene glycol and 1,3-propylene glycol in plasma.
El Amrani M, Zandbergen A, Groot M … +3 more , Bognar T, Smeijsters EH, van der Elst KCM
J Anal Toxicol · 2026 May · PMID 41693000 · Full text
Alcohol and glycol ingestion, including substances such as methanol, ethylene glycol, and isopropanol, constitute a serious medical emergency that requires prompt diagnosis and treatment of patients. Rapid and accurate q...
Alcohol and glycol ingestion, including substances such as methanol, ethylene glycol, and isopropanol, constitute a serious medical emergency that requires prompt diagnosis and treatment of patients. Rapid and accurate quantification of these compounds in plasma is essential to guide clinical decision-making and prevent delays in treatment that could result in irreversible organ damage or death. Many healthcare facilities lack analytical methods capable of simultaneously quantifying both alcohols and glycols in a single run. This study describes the development and validation of a rapid gas chromatography-flame ionization detection (GC-FID) method for the simultaneous screening and quantification of methanol, ethanol, 2-propanol, acetone, ethylene glycol, diethylene glycol, 1,2-propylene glycol, and 1,3-propylene glycol in human plasma. Plasma samples were prepared using a protein precipitation technique with acetonitrile containing two internal standards: 2-butanol and 1,4-butanediol. Acetonitrile effectively precipitated plasma proteins. The supernatant was then subjected to GC-FID analysis for quantification of the target al.ohols and glycols. The total analytical run time was 5 minutes, enabling the quantification of eight analytes in a single injection. The method demonstrated excellent linearity, with correlation coefficients (R2) exceeding 0.9995 for all compounds. The linear dynamic range was 40-1280 mg/L for methanol, 2-propanol, acetone, ethylene glycol, diethylene glycol, 1,2-propylene glycol, and 1,3-propylene glycol, and 80-2560 mg/L for ethanol. Within-run and between-run precision and accuracy (CV and bias) for all analytes were within the predefined acceptance criteria of ±15%. No significant interference, carry-over, or matrix effects were observed, confirming the method's selectivity and robustness. The developed GC-FID method enables rapid, accurate, and simultaneous quantification of toxic alcohols and glycols in plasma within a 5-minute run time. The excellent linearity, precision, and selectivity of the method met al. analytical performance criteria, making it well-suited for routine clinical use. This method provides a valuable tool for timely diagnosis and management of suspected toxic alcohol and glycol ingestions in patients in emergency settings.
The detection of phenibut in keratinized specimens: a validation and case study.
Racines A, Jones JT, Wang M … +1 more , Coy DJ
J Anal Toxicol · 2026 May · PMID 41692998 · Publisher ↗
Phenibut (β-phenyl-γ-butyric acid) is a central nervous system depressant that acts on the GABAB and, to some extent, the GABAA receptors. Phenibut is not approved for human use in the USA or throughout most of Europe du...
Phenibut (β-phenyl-γ-butyric acid) is a central nervous system depressant that acts on the GABAB and, to some extent, the GABAA receptors. Phenibut is not approved for human use in the USA or throughout most of Europe due to its numerous adverse effects, such as intoxication, tachycardia, agitation, and confusion. A method to detect phenibut in hair and nail specimens using solid phase extraction followed by liquid chromatography tandem mass spectrometry was validated. This method was used to analyze a pair of hair and fingernail specimens from a self-reported user of phenibut. The assay validation satisfied all the criteria of the ANSI/ASB. Standard Practices for Method Validation in Forensic Toxicology Standard 036. The measured concentration of phenibut in the hair and the fingernail of the self-reported phenibut user were greater than the upper limit of linearity (>4000 pg/mg). Phenibut continues to be a growing concern in the USA with limited information on public health outcomes due to the scarcity of testing and the interpretation of results. This study is the first report of an analytical method for the detection of phenibut in keratinized specimens, including positive detection in both hair and nail of a self-reported user. Keratinized specimens show promise as alternatives for phenibut detection to the more invasive testing matrices, including urine and blood.
Antidepressant metabolite concentrations and metabolite-to-parent drug ratios in postmortem femoral blood.
Kriikku P, Ojanperä I
J Anal Toxicol · 2026 May · PMID 41692991 · Full text
Antidepressants are commonly used psychiatric medications, many of them associated with significant toxicity in overdose. While established reference values are used in therapeutic monitoring of these drugs, there are le...
Antidepressants are commonly used psychiatric medications, many of them associated with significant toxicity in overdose. While established reference values are used in therapeutic monitoring of these drugs, there are less abundant reference data available in postmortem (PM) toxicology, especially concerning antidepressant metabolites. In this study, all such PM cases in 2016-2023 were retrieved from the Finnish national PM toxicology database in which an antidepressant drug and its metabolite were quantified concurrently in femoral venous blood, representing all causes of death (N = 3472). The results comprise the median concentrations of 11 antidepressants and their main metabolites, the parent drug plus metabolite sum concentrations, and the respective metabolite-to-parent drug ratio (MPR) values. The study revealed that the concentrations of antidepressants and their metabolites in PM blood generally increase in the following order according to the group studied: other cause of death than poisoning < implicated in fatal poisoning < principal finding in fatal antidepressant poisoning < single finding in fatal antidepressant poisoning. MPR was found to be a useful addition to concentration values in distinguishing acute poisoning death from other causes of death, showing lower MPR values in poisoning, and the results obtained in this study compared favorably with data previously published in the literature. The parent drug plus metabolite sum concentration did not provide significant additional information to parent drug only in PM diagnostics of poisoning.
Toxicological interpretation of N-ethylpentedrone and its metabolites in 12 fatal and non-fatal forensic cases.
Roosendaal J, Bosman IJ, Van Driessche PMI … +5 more , Rijken DJ, Hulshof JW, van de Velde B, Klein T, Maudens KEK
J Anal Toxicol · 2026 May · PMID 41663893 · Publisher ↗
N-Ethylpentedrone (NEP) is a synthetic cathinone that remained legally available in the Netherlands until its inclusion under the Opium Act as of 1 July 2025. Although NEP emerged in the mid-2000s, toxicological data, es...
N-Ethylpentedrone (NEP) is a synthetic cathinone that remained legally available in the Netherlands until its inclusion under the Opium Act as of 1 July 2025. Although NEP emerged in the mid-2000s, toxicological data, especially regarding its metabolism and concentration in real-world cases, are scarce. The aim of this study is to demonstrate analytical methods for the detection and quantification of NEP and its metabolites in antemortem and postmortem biological matrices and to provide reference values and guidance for interpretation. Cases where NEP was detected between May 2024 and September 2025 were selected. All cases have undergone Systematic Toxicological Analyses, which consists of a selection of screening procedures (HS-GC-FID and LC-QToF-MS) and a targeted semi-quantitative LC-MS/MS method. Additional quantitative LC-MS/MS analyses were performed if needed. NEP metabolites were investigated by LC-Q-Orbitrap-MS. NEP was detected in 12 cases (all male, age range 18-56 years). The median antemortem (n = 8) and postmortem (n = 5) NEP (femoral) blood concentrations were 0.062 mg/L (range <0.005-0.45 mg/L) and 0.63 mg/L (range 0.26-0.93 mg/L), respectively. In four cases, a NEP (multidrug) intoxication was concluded as a possible cause of death. One case indicated a half-life of around 2-3 hours, a blood/plasma ratio of around 1.1 and an up to 5-fold postmortem increase of NEP within 48 hours after death. Several phase-1 metabolites of NEP were detected, but only NEP itself was identified in all investigated blood, plasma, urine, and vitreous humor samples. NEP recently resurfaced in the Netherlands, with four potentially fatal cases. For the analysis and interpretation of NEP blood concentrations, degradation of NEP in blood stored at room temperature, (survival) time since intake and a potential significant postmortem increase should be taken into account. NEP itself remains the target compound of choice, with dihydro-NEP and N-desethyl-NEP serving as complementary analytes depending on case circumstances.
Micro-QuEChERS extraction method optimization for quantification of designer benzodiazepines in forensic postmortem blood samples.
Souza KAO, Teodoro JAR, Berlinck DZ … +7 more , Rezendes APK, de Paula DML, Souza Pelição F, de Aquino EM, Gianvecchio VAP, Yonamine M, Costa JL
J Anal Toxicol · 2026 May · PMID 41652886 · Publisher ↗
The aim of this study was to develop and validate a quantitative method for the analysis of designer benzodiazepines in postmortem blood samples using micro-QuEChERS extraction and liquid chromatography tandem mass spect...
The aim of this study was to develop and validate a quantitative method for the analysis of designer benzodiazepines in postmortem blood samples using micro-QuEChERS extraction and liquid chromatography tandem mass spectrometry (LC-MS/MS). A comprehensive optimization of the method was performed using a multivariate statistical approach, incorporating validation criteria in line with established practices for method validation in forensic toxicology. The method showed linearity between 1 and 200 ng/mL (r2 > 0.990), with good imprecision (<9.8%) and inaccuracy (<11.1%) evaluated at three different quality control concentrations. Matrix effects and recovery rates were found to be better than 58% and 77.5%, respectively. No carryover or interferences were detected during the analysis. The method was effectively utilized on two real forensic postmortem blood samples, both of which tested positive for bromazolam, showing concentrations of 31 and 40 ng/mL. The micro-QuEChERS extraction method demonstrated satisfactory analytical performance and is an environmentally sustainable option, minimizing the use of solvents and reagents, with potential for application in both clinical and forensic analyses, aligning with green analytical toxicology principles.
False positive urine drug screens.
Saitman A, Fitzgerald RL, Lund K … +2 more , Suhandynata RT, Menlyadiev M
J Anal Toxicol · 2026 May · PMID 41639014 · Publisher ↗
Immunoassay-based urine drug screens are widely employed in clinical toxicology due to their speed, low cost, and ease of automation. However, these assays are inherently limited by antibody cross-reactivity, which can r...
Immunoassay-based urine drug screens are widely employed in clinical toxicology due to their speed, low cost, and ease of automation. However, these assays are inherently limited by antibody cross-reactivity, which can result in false-positive findings and incorrect interpretations with major implications for patient care, employment, and legal outcomes. This review updates prior literature by analyzing reported false-positive interferences published between 2013 and 2024 across commonly screened drug classes, including opioids, amphetamines, benzodiazepines, cannabinoids, barbiturates, phencyclidine (PCP), cocaine, ethanol, and ethyl glucuronide. A total of 61 studies met inclusion criteria from 569 unique publications retrieved via PubMed. Each report was categorized by level of evidence, ranging from single case reports to controlled spiking experiments. Despite advances in antibody specificity, immunoassay drug screens remain presumptive and require confirmation by orthogonal techniques such as gas or liquid chromatography coupled with mass spectrometry (GC-MS or LC-MS/MS). This review provides updated reference data on known interferents, emphasizes the need for laboratorian-clinician communication, and supports continued education on assay limitations. Reliable interpretation of presumptive immunoassay drug screen results remains essential to prevent inappropriate clinical care decisions.
Further elucidation of the metabolism of 2,4-dinitrophenol in a case of unexpected death.
Vanthourenhout S, Deprez C, Maes P … +5 more , Carlier L, Van Hoorick M, Heylen O, Mulliez S, Croes K
J Anal Toxicol · 2026 Apr · PMID 41530400 · Publisher ↗
As a weight-loss drug, 2,4-dinitrophenol (2,4-DNP) was discovered and popularized in the 1930s. Owing to multiple adverse effects, including death, its use as a prescription drug was banned in 1938. However, toxicity cas...
As a weight-loss drug, 2,4-dinitrophenol (2,4-DNP) was discovered and popularized in the 1930s. Owing to multiple adverse effects, including death, its use as a prescription drug was banned in 1938. However, toxicity cases have risen over the past two decades as 2,4-DNP is easily obtained illegally online. In a fatal case at our hospital involving rapid, unexplained deterioration, 2,4-DNP was identified through a full toxicological analysis of blood and urine. An LC-MS/MS method for detecting and quantifying 2,4-DNP in plasma and urine was developed and validated according to European Medicines Agency (EMA) criteria. It also allowed quantification of its main metabolites 2-amino-4-nitrophenol (2A-4NP) and 4-amino-2-nitrophenol (4A-2NP) in urine. In plasma, 2A-4NP was only semi-quantifiable; 4A-2NP was undetected, likely due to matrix effects reducing sensitivity. Results obtained with and without enzymatic hydrolysis showed that 2,4-DNP-glucuronide and 2A-4NP plus its glucuronide are the primary metabolites, whereas 4A-2NP and its glucuronide are less prominent. No sulfate conjugates were detected. This study is the first to compare sample preparation with and without enzymatic hydrolysis, offering new insights into 2,4-DNP metabolism and the relative importance of its major metabolites and their glucuronides.
Validation of a microsampling-compatible liquid-liquid extraction method for cannabinoid quantitation in 50 µL of whole blood using liquid chromatography-mass spectrometry.
Mohammed AA, Khan M, Chan H … +1 more , Brubacher JR
J Anal Toxicol · 2026 Apr · PMID 41520150 · Full text
Recently developed dried blood analysis methods for cannabinoid quantitation utilize small blood volumes, making them microsampling-compatible, but are limited by hematocrit-related bias for dried blood spots (DBSs) and...
Recently developed dried blood analysis methods for cannabinoid quantitation utilize small blood volumes, making them microsampling-compatible, but are limited by hematocrit-related bias for dried blood spots (DBSs) and higher consumable costs for volumetric absorptive microsampling (VAMS®). To address these issues, we developed a highly sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method capable of quantifying cannabinoids in 50 µL of liquid whole blood, providing a practical microsampling alternative to dried blood approaches. Using liquid-liquid extraction (LLE) with sodium hydroxide alkalinization and acetonitrile precipitation, followed by quantitative analysis on an Agilent 6495 liquid chromatography-triple quadrupole mass spectrometer, we achieved lower limits of quantitation (LLOQ) of 0.10 ng/mL for Δ9-tetrahydrocannabinol (THC), cannabinol (CBN), and cannabigerol (CBG), 0.20 ng/mL for cannabidiol (CBD), 0.50 ng/mL for 11-hydroxy-THC (11-OH-THC), and 1.0 ng/mL for 11-nor-9-carboxy-THC (THC-COOH). Calibration was linear from the LLOQ to 300 ng/mL for all analytes. To our knowledge, this is one of the first validated LLE approaches for cannabinoid quantitation in less than 100 µL of liquid whole blood. Further, it achieves sub-ng/mL sensitivity, exceeding the LLOQs of most published methods which require ≥100 µL of whole blood. We anticipate particular utility for our method in obtaining evidence from suspected impaired drivers at the roadside when paired with capillary microsampling, such as via finger prick. This approach enables measurement of THC levels at the time of driving, thereby overcoming current limitations, including the decrease in THC levels that occurs with delayed blood sampling, requirement for larger sample volumes (≥100 µL), and dependence on trained phlebotomists for venipuncture.
Malicious poisoning of a canine with the neuromuscular blocking agent rocuronium: a case report with postmortem quantitation.
Buchweitz JP, Zyskowski JA, Cajigas K … +1 more , Brooks JW
J Anal Toxicol · 2026 Jun · PMID 41514164 · Full text
Rocuronium bromide is a non-depolarizing neuromuscular blocking agent (NMBA) commonly used in anesthesia for its rapid onset and intermediate duration of action. While its therapeutic use has been well-documented in huma...
Rocuronium bromide is a non-depolarizing neuromuscular blocking agent (NMBA) commonly used in anesthesia for its rapid onset and intermediate duration of action. While its therapeutic use has been well-documented in humans and experimentally in dogs, this report describes the first known case of its malicious use in the fatal intoxication of a canine. Herein we describe the case of a six-year-old male Plott Hound found deceased under suspicious circumstances. Postmortem investigation revealed focal hemorrhages near potential injection sites, and toxicological analysis via liquid chromatography-tandem mass spectrometry confirmed the presence of rocuronium in muscle tissue, heart blood, and urine. The dog had no surgical history or indication for anesthesia. However, rocuronium concentrations mirrored those seen in anesthetized human patients under ventilatory support. The owner, a licensed pharmacist, was suspected of drug theft and aggravated animal abuse. This case highlights the lethality of NMBAs when misused outside of the clinical setting and further underscores a collaborative, multi-institutional, multidisciplinary approach to veterinary forensic investigations.
Adsorption of Tetrahydrocannabinol, Metabolites, and Related Cannabinoids During Storage of Plasma Samples in Gel Separation Tubes.
Abdul IA, Pon D, Giancola C … +1 more , Woodall K
J Anal Toxicol · 2026 Mar · PMID 41496001 · Publisher ↗
The concentrations of some drugs in biofluids can be affected by different storage conditions, including the type of sample collection tube. This phenomenon has been observed in gel separation tubes, where drug adsorptio...
The concentrations of some drugs in biofluids can be affected by different storage conditions, including the type of sample collection tube. This phenomenon has been observed in gel separation tubes, where drug adsorption to the gel separator can lead to the underestimation of the drugs concentration, thus potentially affecting the interpretation of analytical results. The purpose of this study was to determine if concentrations of tetrahydrocannabinol (THC), its metabolites, and related cannabinoids decrease over time when stored in plasma separation tubes (PSTs) as compared to non-PSTs. Plasma samples with a high concentration [HP-24 ng/mL THC, 150 ng/mL carboxy-THC (THC-COOH), 48 ng/mL hydroxy-THC (THC-OH), 15 ng/mL cannabidiol (CBD), and 15 ng/mL cannabinol (CBN)], and low concentration [LP-5.0 ng/mL THC, 31 ng/mL THC-COOH, 10 ng/mL THC-OH, 3.1 ng/mL CBD, and 3.1 ng/mL CBN] of cannabinoids were stored in PSTs and non-PSTs for analysis by liquid chromatography-tandem mass spectrometry at one-hour, three-day, one-week, two-week, three-week, one-month, two-month, and three-month intervals. Statistically significant differences in cannabinoid concentrations (p < .05) were observed between non-PSTs and PSTs. All cannabinoids except THC-COOH showed a greater reduction in concentration when stored in PSTs compared to non-PSTs. In contrast, THC-COOH showed an increase in concentration when stored in PSTs compared to non-PSTs. Over a three-month period, concentrations in PSTs decreased for THC by 83% and 81%, THC-OH by 66% and 63%, CBD by 69% and 62%, and CBN by 75% and 70% in LP and HP samples, respectively. In conclusion, for forensic cases involving cannabinoids, the adsorption of these compounds should be considered in the toxicological interpretation of samples collected in PSTs.
Aldehyde dehydrogenase 2 and sex influence blood acetaldehyde levels in mice, but not ethanol levels.
Jamal M, Takei S, Miki T … +3 more , Tsukamoto I, Kinoshita H, Murase T
J Anal Toxicol · 2026 May · PMID 41495998 · Publisher ↗
This study measured the concentrations of blood ethanol (EtOH) and acetaldehyde (AcH) in mice to examine the roles of aldehyde dehydrogenase 2 (ALDH2) and sex following intragastric administration of EtOH. The experiment...
This study measured the concentrations of blood ethanol (EtOH) and acetaldehyde (AcH) in mice to examine the roles of aldehyde dehydrogenase 2 (ALDH2) and sex following intragastric administration of EtOH. The experiment utilized males and females of two mouse strains: C57BL/6N (wild-type, WT) and Aldh2-knockout (Aldh2-KO) mice. Aldh2-KO mice lack the ALDH2 enzyme, leading to the accumulation of high levels of AcH in the blood. The mice were fasted for approximately six hours before EtOH administration. EtOH (1.0, 2.0, and 3.0 g/kg) was administered intragastrically, and blood samples were collected at 30, 60, 120, 180, 240, and 300 minutes post-EtOH administration through retro-orbital puncture. The samples were then analyzed using headspace gas chromatography. The results for both male and female WT mice showed that EtOH and AcH levels increased in a dose-dependent manner, peaked at 60 minutes post-ingestion, and then gradually decreased. While there were no significant differences in blood EtOH concentrations between males and females, the concentrations of AcH were significantly higher in female mice than in male mice, indicating potential sex-related differences in EtOH metabolism. In Aldh2-KO mice, the EtOH and AcH levels increased initially and peaked at 30-60 minutes post-ingestion, with no significant differences in EtOH or AcH concentrations between the sexes. While the concentrations of EtOH in both male and female Aldh2-KO mice gradually decreased, the concentration of AcH remained elevated until six hours post-ingestion due to the ALDH2 deficiency inhibiting AcH oxidation. Our findings emphasize the importance of considering the influences of sex and ALDH2 when researching the effects of alcohol, particularly in relation to the EtOH byproduct AcH.
4-Anilino-N-phenylethylpiperidine (4-ANPP): the potential caution flag for illicit fentanyl.
Laraia N, Bierly JJ, Chan-Hosokawa A
J Anal Toxicol · 2026 Apr · PMID 41495478 · Publisher ↗
The ability to distinguish illicit fentanyl use is becoming increasingly critical in toxicological investigations. 4-Anilino-N-phenylethylpiperidine (4-ANPP), also known as despropionylfentanyl, is both a precursor in il...
The ability to distinguish illicit fentanyl use is becoming increasingly critical in toxicological investigations. 4-Anilino-N-phenylethylpiperidine (4-ANPP), also known as despropionylfentanyl, is both a precursor in illicit fentanyl production and a minor metabolite frequently detected alongside fentanyl in forensic toxicology. Its presence may assist in distinguishing medical and illicit fentanyl sources. This study evaluated 4-ANPP concentrations and 4-ANPP: fentanyl (4-ANPP to fentanyl) ratios in clinical (presumed medicinal) and postmortem (forensic) submissions to ascertain trends that may aid in source attribution and toxicological interpretation. Blood and serum/plasma (s/p) samples were analyzed via liquid-liquid extraction followed by liquid chromatography tandem mass spectrometry (LC-MS/MS) throughout 2023. A total of 32 723 forensic and 1015 clinical cases positive for fentanyl were included in the analysis. Most clinical 4-ANPP concentrations (69%) were below 0.50 ng/mL, compared to 20% of forensic cases. Forensic blood samples with reportable 4-ANPP concentrations (n = 29 701) had a median of 2.1 ng/mL (mean ± SD: 6.5 ± 42 ng/mL, range: 0.20-4100 ng/mL). In clinical serum/plasma samples with reportable 4-ANPP (n = 451), the median was 0.96 ng/mL (mean ± SD: 3.9 ± 17 ng/mL, range: 0.20-306 ng/mL). In cases with reportable 4-ANPP concentrations, the median 4-ANPP: fentanyl ratio was 0.141 (mean ± SD: 0.22 ± 0.95; range: 0.000078-140) for forensic, while the clinical median was 0.105 (mean ± SD: 0.44 ± 3.0; range: 0.005-60). Notably, 91% of forensic cases had reportable 4-ANPP concentrations (≥0.20 ng/mL) compared to 44% of clinical cases, excluding more than half of the clinical cases from ratio calculations. Although overlapping 4-ANPP: fentanyl ratios limit its utility as a clear indicator of illicit fentanyl use, elevated 4-ANPP concentrations are more strongly associated with non-pharmaceutical sources and may serve as valuable support in forensic interpretation.
Letter to the editor concerning the article: Aknouche F, Magny R, Maruejouls C, Trebuchet C, Fargeot K, Thion L, Valancony C, Scherrer F, Oussedik ND, Kintz P, Labat L, Houzé, P. (2025) Systemic organophosphate poisoning in child following anti-lice lotion application. Journal of Analytical Toxicology, bkaf096.
Dona A
J Anal Toxicol · 2026 Apr · PMID 41495452 · Publisher ↗
Abstract loading — click title to view on PubMed.
About
- Frequency
- Sun
- Papers found
- 200
- RSS feed
- Subscribe