Journal Of Analytical Toxicology[JOURNAL]
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200 papers
Recent trends in the analysis of GHB in hair.
Kustera C, LeBeau M, Sharma S … +1 more , Arroyo L
J Anal Toxicol · 2025 Nov · PMID 40676726 · Publisher ↗
Hair analysis is a valuable tool in forensic toxicology, providing extended detection windows and critical insights into drug testing, usage trends, and drug-facilitated crimes. This systematic review was conducted using...
Hair analysis is a valuable tool in forensic toxicology, providing extended detection windows and critical insights into drug testing, usage trends, and drug-facilitated crimes. This systematic review was conducted using Scopus, Web of Science, and PubMed databases from March 2017 to September 2024, and evaluated 19 studies (16 research articles and 3 case reports) on the detection of γ-hydroxybutyrate (GHB) in hair. This review examines recent studies on GHB concentrations in hair, focusing on both endogenous and exogenous concentrations resulting from illicit and prescribed use, as well as the analytical methods employed. This review includes decontamination parameters, extraction techniques, and sample sizes used during the analytical method. New studies report that endogenous GHB levels range from 0.2 to 5.5 ng/mg, while exogenous levels vary widely from 0.3 to 239.6 ng/mg. Additionally, published results indicate that the frequency of use may be more significant than the dosage for exogenous GHB to be incorporated into the hair. A novel adjacent segmentation method has been proposed to differentiate endogenous from exogenous GHB, identifying local peaks within adjacent hair segments. Research into GHB-glucuronide as a biomarker has found it unreliable due to inconsistent correlations with exogenous use. Further research is needed to refine the interpretation of GHB levels in forensic applications.
Chlorthalidone in vitro metabolite identification for documenting exposure in doping.
Di Giorgi A, Daziani G, Tini A … +3 more , Tronconi L, Carlier J, Taoussi O
J Anal Toxicol · 2026 Apr · PMID 40668262 · Publisher ↗
Diuretics are commonly used in doping because they can conceal the presence of performance-enhancing substances in an athlete's urine through dilution and promote rapid weight loss. As a result, these substances are proh...
Diuretics are commonly used in doping because they can conceal the presence of performance-enhancing substances in an athlete's urine through dilution and promote rapid weight loss. As a result, these substances are prohibited in sports by the World Anti-Doping Agency (WADA) under the S5 category ("Diuretics and Masking Agents"). Chlorthalidone, a thiazide-like diuretic, is medically used as an antihypertensive agent and is prescribed for conditions such as heart failure and liver cirrhosis. However, it is also misused in doping. The detection of chlorthalidone or its metabolite markers in an athlete's urine is essential to prove consumption. Therefore, the aim of the study was to assess the metabolism of the substance in humans. For this purpose, chlorthalidone metabolites were predicted with GLORYx (Hamburg University, Germany) to identify the transformations that may occur with higher probability; the compound was incubated with 10-donor-pooled human hepatocytes to simulate hepatic metabolism; and the incubates were analyzed by liquid chromatography-high-resolution tandem mass spectrometry (LC-HRMS/MS) and software-aided data mining. In silico simulations predicted 11 Phase II metabolites, with N-acetylation at the sulfonamide group being the predominant transformation (88% probability score); other major reactions included O-glucuronidation, O-sulfation, and glutathione conjugation, with probability scores lower than 70%. Two metabolites were identified in in vitro hepatocyte incubates and presented a reduction or a hydroxylation at the phthalimidine moiety. To the best of the authors' knowledge, these metabolites are specific to chlorthalidone and can be targeted as markers for analytical screening in anti-doping controls.
Artificial intelligence in new psychoactive substances analysis: state-of-art and future perspectives.
Di Giorgi A, Pichini S, Busardò FP … +1 more , Basile G
J Anal Toxicol · 2026 Apr · PMID 40668255 · Publisher ↗
Analytical toxicology is a discipline of forensic toxicology which applies analytical techniques for the determination of drugs of abuse in biological and nonbiological matrices. To this concern, artificial intelligence...
Analytical toxicology is a discipline of forensic toxicology which applies analytical techniques for the determination of drugs of abuse in biological and nonbiological matrices. To this concern, artificial intelligence (AI), particularly machine learning (ML), is innovating analytical toxicology by improving data processing and facilitating the identification of New Psychoactive Substances (NPS). The aim of this review was to explore the current application of AI in this field and to highlight the future perspectives. A literature search was performed in several scientific databases to review articles reporting the implementation of AI models for analytical toxicological purposes. The most frequent applications of these technologies were for compound identification, molecular structure prediction and retention time prediction. AI proved to be a valuable tool for analytical toxicologists for the capability to process large amount of data which are typically obtained by untargeted approaches.
Development and application of an LC-MS/MS method for 8 antiepileptic drugs and 2 metabolites using microsampling techniques (DBS and VAMS).
Cobo-Golpe M, Paniagua-González L, Lendoiro E … +12 more , Blanco-Ces M, López-Rabuñal Á, Abella J, Castro D, Melcón C, Fuentes P, Carballeira I, García C, Gómez C, López-Rivadulla Lamas M, Cruz A, de-Castro-Ríos A
J Anal Toxicol · 2026 Apr · PMID 40668253 · Full text
Therapeutic drug monitoring (TDM) of antiepileptic drugs (AEDs) is used for optimization and individualization of patients' treatment. Capillary microsampling techniques are a promising alternative to conventional venous...
Therapeutic drug monitoring (TDM) of antiepileptic drugs (AEDs) is used for optimization and individualization of patients' treatment. Capillary microsampling techniques are a promising alternative to conventional venous sampling for TDM. Both dried blood spots (DBS) and volumetric adsorptive microsampling (VAMS) devices are less invasive and patient-friendly sampling techniques which have been gaining interest in the last years. This study describes the development and validation of an LC-MS/MS method for the determination of 8 AEDs (Carbamazepine, Lacosamide, Levetiracetam, Lamotrigine, Phenobarbital, Valproic acid) and 2 metabolites (10,11-Dihydro-10-hydroxy-carbamazepine (DHCB) and carbamazepine-10,11-epoxide) in DBS and VAMS samples. The method was fully validated for linearity, selectivity, accuracy, precision, carryover, matrix effect, recovery and stability (15 days at room temperature and 72 h in autosampler). Moreover, the volume effect, volcano effect, and the hematocrit (Hct) effect were also assessed for DBS samples. All parameters showed satisfactory results, with a limit of quantification ranging from 0.5 to 10 µg/mL, depending on the analyte. Some instability issues were detected in DBS samples for oxcarbazepine. However, the inclusion of oxcarbazepine's metabolite DHCB overcomes this problem as it was stable under both conditions tested. Moreover, this is the first DBS or VAMS method reporting the inclusion of DHCB, which seems essential for the TDM of oxcarbazepine. The method was applied to 80 paired samples from patients under treatment with these drugs in order to study the suitability of the method for the detection of these compounds, and compare concentrations in paired VAMS, DBS, whole blood and plasma samples. Ratios between paired samples show a promising correlation between microsampling techniques and plasma concentrations.
Supporting evidence for 5-acetamido-N-desethylmetonitazene as a potential metabolite rather than a degradation product.
Wood R, Moore R
J Anal Toxicol · 2025 Sep · PMID 40665518 · Publisher ↗
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Prevalence and concentration of fentanyl in hair collected for court-ordered mandatory drug testing.
Grabenauer M, Bynum ND, Johann LE … +5 more , Bollinger K, Davis LS, Hayes ED, Flegel RR, Winecker RE
J Anal Toxicol · 2025 Nov · PMID 40663381 · Publisher ↗
Hair testing is often employed by court-ordered mandatory drug testing (COMDT) programs; however, as of December 2024, many of these programs still do not include fentanyl in their testing panels. Further, testing panels...
Hair testing is often employed by court-ordered mandatory drug testing (COMDT) programs; however, as of December 2024, many of these programs still do not include fentanyl in their testing panels. Further, testing panels including fentanyl for purposes of workplace testing are rare, and concentrations of fentanyl in hair of people who have used drugs are needed to validate future testing cutoffs. In this study, we analyzed 1025 hair specimens, originally collected for COMDT purposes, for 26 substances, including 13 fentanyl-related compounds. Methamphetamine was the most frequently detected compound (n = 266, 26%), followed by hydrocodone (n = 157, 15%). Fentanyl was the most detected fentanyl-related compound, followed by 4-ANPP. Fentanyl was detected in 151 (15%) hair specimens. 12 specimens contained a fentanyl-related compound with no detectable fentanyl. Of the 163 specimens in which fentanyl or a fentanyl-related compound was detected 31 (19%) had no other analytes detected. Using a cutoff of 1 pg/mg the detection rate for fentanyl was 14.7%. Conversely, most commercial testing laboratories utilize cutoffs between 20 and 100 pg/mg. For the 98 specimens with fentanyl concentrations in the quantifiable range (5-2000 pg/mg), the maximum, mean, and median concentrations were 1946, 223, and 55 pg/mg, respectively. An additional 7 specimens had concentrations greater than the upper limit of quantification of 2000 pg/mg with an estimated maximum fentanyl concentration of 9246 pg/mg. Forty-four specimens contained detectable norfentanyl. The norfentanyl: fentanyl ratios ranged from 0.02 to 0.46 with a mean of 0.09. COMDT programs that do not include fentanyl or employ common commercial cutoffs in their testing protocols for fentanyl are potentially missing drug positive specimens.
Balancing the (uncertainty) budget-postmortem toxicology laboratory conformance to ANSI/ASB Standard 056: Standard for Evaluation of Measurement Uncertainty in Forensic Toxicology.
Kahl JH, Moore DM
J Anal Toxicol · 2025 Nov · PMID 40660623 · Publisher ↗
In postmortem forensic toxicology, the accuracy and reliability of toxicological results are critical to the medicolegal death investigation process. ANSI/ASB Standard 056: Standard for Evaluation of Measurement Uncertai...
In postmortem forensic toxicology, the accuracy and reliability of toxicological results are critical to the medicolegal death investigation process. ANSI/ASB Standard 056: Standard for Evaluation of Measurement Uncertainty in Forensic Toxicology establishes the minimum requirements for evaluating measurement uncertainty (MU) in quantitative methods utilized in forensic toxicology. Accurate evaluation of MU increases confidence in results, supports scientific rigor, enables inter-laboratory comparability, and ensures legal defensibility. Using the National Institute of Standards and Technology (NIST) 8-step procedure described in ANSI/ASB Standard 056, postmortem forensic toxicology laboratories can develop customized, flexible MU budget templates that accommodate a variety of analytical workflows and sample preparation techniques commonly used in the field. This manuscript highlights the use of a template that is adaptable to both routine quantitative workflows and those employing method of standard addition, providing example MU calculations for each. By aligning laboratory practices with the NIST 8-step procedure, as well as integrating accreditation requirements and published ANSI/ASB Standards into their quality management system, laboratories enhance the accuracy and reliability of their toxicological results. Adhering to ANSI/ASB Standard 056 ensures that the inherent variability in postmortem toxicological analyses is appropriately assessed and managed in a manner consistent with best practices.
The detection of cannabinoids in breath after ingestion of cannabis-infused edibles.
Bery JL, Brooks-Russell A, Lovestead TM … +1 more , Jeerage KM
J Anal Toxicol · 2025 Nov · PMID 40638229 · Full text
The increase of Δ9-tetrahydrocannabinol (THC) in breath after cannabis inhalation has been well-documented in the forensic field, but the trends after ingestion of cannabis-infused edibles have not yet been investigated....
The increase of Δ9-tetrahydrocannabinol (THC) in breath after cannabis inhalation has been well-documented in the forensic field, but the trends after ingestion of cannabis-infused edibles have not yet been investigated. In this study, participants ingested a cannabis-infused edible and provided breath samples before and at three timepoints after ingestion. Participants were assigned to one of two breath sampling devices. THC was found in most pre-use breath samples, and THC concentration had variable trends after ingestion. Nineteen participants exhibited a maximum in their THC concentration at 47, 92, or 180 min after ingestion, while six participants had their highest THC concentration before the observed ingestion, and four participants had no significant change in THC concentration over the four samples. Five additional cannabinoids were detected in breath. While cannabidiol (CBD) trends followed THC trends for some participants, diverging trends in other participants suggest different biological processing of CBD derived from edibles. This proof-of-concept study shows that THC concentration in breath can increase after the ingestion of cannabis-infused edibles, but the uncertainty of breath measurements and a longer time window need to be further explored.
Fatalities following DMT use: two case reports and a review of the literature.
Pullen J, Moore R, Wood R … +3 more , Rab E, Couchman L, Copeland CS
J Anal Toxicol · 2025 Nov · PMID 40632616 · Full text
N, N-Dimethyltryptamine (DMT) is a hallucinogen found in the South American Psychotria viridis plant and is the major psychoactive ingredient in the brew ayahuasca. In this report, we performed a review of the surroundin...
N, N-Dimethyltryptamine (DMT) is a hallucinogen found in the South American Psychotria viridis plant and is the major psychoactive ingredient in the brew ayahuasca. In this report, we performed a review of the surrounding literature and detail two deaths which recently occurred in the UK following DMT use. A literature search of both academic (PubMed, GoogleScholar) and media (using Google search engine) publications was performed to identify previously reported fatalities following DMT use. The National Programme on Substance Use Mortality (NPSUM) was also searched for deaths which have occurred in the UK following DMT use. Literature search-There have been three previous reports of fatalities following DMT use, all deemed accidental in nature, with DMT consumption taking place as part of an ayahuasca ceremony in two of these cases. NPSUM cases-Two cases were identified (Case Report 1 [CR1] & Case Report 2 [CR2]), neither of which occurred in the context of an ayahuasca ceremony. DMT was detected and quantified in femoral blood in both cases (CR1 0.23 mg/l; CR2 0.24 mg/l). There was evidence of polydrug use in both cases (CR1 n = 6; CR2 n = 9), which in each case included additional compounds which can increase serotonergic drive (CR1 cocaine, amphetamine; CR2 venlafaxine, mirtazapine). There have been two recent deaths following DMT use in the UK, both in the context of polydrug use which may have caused death due to excessive serotonergic innervation leading to serotonin syndrome. Polydrug use is increasingly common in the UK, and users of unregulated drugs should caution their use in combination with other unregulated drugs and also any prescribed medications.
Exploring transdermal SARMs exposure: analysis of the elimination profiles and metabolism for doping control purposes.
Korsmeier L, Krombholz S, Alhalabi H … +2 more , Thomas A, Thevis M
J Anal Toxicol · 2025 Nov · PMID 40632609 · Publisher ↗
Transdermal drug delivery has been of particular interest to pharmaceutical research for decades, but is also becoming increasingly relevant in the field of sports drug testing. As shown in the past, the (unintentional)...
Transdermal drug delivery has been of particular interest to pharmaceutical research for decades, but is also becoming increasingly relevant in the field of sports drug testing. As shown in the past, the (unintentional) oral ingestion of trace amounts of prohibited selective androgen receptor modulators (SARMs), e.g. due to product contamination, can lead to an adverse analytical finding (AAF) in doping controls. Another site of exposure is presented by the skin, as it provides a large surface for drug penetration. However, the extent of diffusion through various layers of the skin and into the blood vessels depends, among other things, on the physicochemical and biological properties of a substance. The objective of this project was to simulate a transdermal contamination scenario and investigate the skin penetration and subsequent metabolism of microdoses of three commonly used SARMs: LGD-4033, RAD140, and S-23. For this purpose, an administration study was conducted, in which either 10 or 50 µg of the substances were applied to the lower forearm of 5 volunteers each. The collected urine samples were analyzed via LC-MS/MS following enzymatic hydrolysis and solid-phase extraction. This methodical approach is distinguished by its high sensitivity, enabling the detection of at least 5 pg/mL for LGD-4033 and S-23. After 10 µg administration, LGD-4033 and S-23 as well as associated metabolites were detected, while RAD140 was only detected in urine samples of one subject (n = 5). Following the application of 50 µg, RAD140 was detected in all subjects (n = 5) for up to 9 days, and additional metabolites of LGD-4033 and S-23 were identified. The long-term metabolite of LGD-4033 (M5b) was detected up to 12 days after the dermal administration of 10 µg, and up to 25 days after application of 50 µg, while S-23 was traceable for up to 16, respectively 24 days. It was demonstrated for all three SARMs that they penetrate the skin and may-even in trace amounts-produce AAFs when administered transdermally. Information on urinary concentrations and metabolism following transdermal administration of SARMs may assist in the interpretation of AAFs, particularly when dermal contamination or intentional doping via the skin is discussed.
Evaluation of the iScreen™ urine test FUO drug screen cup for detection of 17 drugs of abuse in urine.
Goebl JA, Zhao F, Zhong J … +2 more , Green C, Han S
J Anal Toxicol · 2025 Nov · PMID 40632604 · Publisher ↗
Drug overdoses are among the most common causes of death in the United States, with synthetic opioids such as fentanyl implicated in the majority of overdose fatalities in the last 10 years. As such, effective rapid assa...
Drug overdoses are among the most common causes of death in the United States, with synthetic opioids such as fentanyl implicated in the majority of overdose fatalities in the last 10 years. As such, effective rapid assays capable of screening against large drugs of abuse panels that include synthetic opioids are critical tools for detecting drug abuse. The iScreen™ Urine Test FUO Drug Screen Cup (Abbott Laboratories) is a multiplexed lateral flow device designed for the preliminary qualitative screening drugs of abuse in urine for forensic applications, and can be used to simultaneously screen urine specimens for up to 17 different drugs of abuse, with multiple potential configurations of assays and cutoffs to support 22 different assay/cutoff combinations. This investigation focused on evaluating the performance of the iScreen™ Urine Test FUO Drug Screen Cup for analytical sensitivity, cross-reactivity characteristics for structurally-related compounds, and method comparison versus the gold standard method (GC-MS or LC-MS/MS). Analytical sensitivity testing demonstrated ≥95% accuracy for all 22 assays during evaluation against both negative and 3x cutoff positive control specimens, and all 22 assays achieved ≥89% concordance with the established reference methodologies in testing with positive and negative human urine specimens.
Application of professional best practices in postmortem forensic toxicology.
Truver MT, Chronister CW, Davis GG … +14 more , Gray TR, Hartman RL, Kahl JH, Karschner EL, Kerrigan S, Kronstrand R, Krotulski AJ, Lee D, Logan BK, Moore DM, Rodda LN, Vikingsson S, Winecker RE, Goldberger BA
J Anal Toxicol · 2025 Oct · PMID 40632588 · Publisher ↗
Postmortem forensic toxicology plays a critical role in medicolegal death investigations through the identification and quantitation of drugs and other substances in postmortem fluids and tissues. Due to the complexity o...
Postmortem forensic toxicology plays a critical role in medicolegal death investigations through the identification and quantitation of drugs and other substances in postmortem fluids and tissues. Due to the complexity of this sub-discipline, consistent application of best practices is critical for ensuring accurate and reliable results, particularly in the context of challenges such as emerging novel psychoactive substances, complex poly-drug interactions, postmortem drug redistribution, and analytical limitations inherent with postmortem specimens. Although there has been significant progress in the development of consensus-based forensic toxicology standards, their scope is intentionally broad to accommodate human performance, postmortem, regulated and non-regulated employment drug testing, court-ordered toxicology, and other applications. Consequently, some aspects specific to postmortem toxicology and medicolegal death investigation are not addressed within the standards. This manuscript seeks to fill these gaps by demonstrating how current standards can be applied in a postmortem toxicology setting and presenting best practices in situations where no established standards exist. These best practices will aid laboratories in prioritizing changes to workflows, allocating resources more efficiently, improving analytical accuracy and reproducibility, ensuring interpretative consistency, and strengthening forensic defensibility in administrative and legal proceedings. Key topics addressed include specimen collection and case submission protocols, method validation approaches tailored for postmortem analysis, optimized analytical workflows based on testing scope and case classification, and quality assurance requirements. Considerations for data review, reporting, and result interpretation are discussed in the context of accurate determination of cause and manner of death. Emphasis is placed on integrating toxicological findings with investigative and autopsy information obtained through ongoing communication with stakeholders. By integrating the application of existing consensus standards with the best community practices for postmortem toxicology, this manuscript aims to support the generation of robust and reliable toxicological data, with the goal of improving forensic investigations, public health surveillance, and drug policy development.
A new automated method for the analysis of cotinine and trans-3'-hydroxycotinine in serum by LC-MS/MS.
Hopkins DL, Weaver ML, Sosnoff C … +3 more , Ahamed R, Wang L, Seyler TH
J Anal Toxicol · 2025 Nov · PMID 40608970 · Full text
Tobacco cigarette smoking is the leading cause of preventable diseases and death in the USA. Exposure to secondhand smoke (SHS) can also cause heart disease, lung cancer, and respiratory illness. Cotinine (COT) and trans...
Tobacco cigarette smoking is the leading cause of preventable diseases and death in the USA. Exposure to secondhand smoke (SHS) can also cause heart disease, lung cancer, and respiratory illness. Cotinine (COT) and trans-3'-hydroxycotinine (HCT) are the primary metabolites of nicotine, the main addictive alkaloid in tobacco products. For many years, we have measured serum levels of COT and HCT in National Health and Nutritional Examination Survey (NHANES) participants to monitor exposure of the US population to active smoking and SHS. As exposure to SHS is decreasing, a more sensitive analytical method is needed to detect the lower levels of these biomarkers for SHS assessment. We developed and validated a new automated method for the detection of COT and HCT in human serum. We implemented a new liquid handling automation system to aliquot and prepare samples using supported liquid extraction. Samples were analyzed by liquid chromatography-tandem mass spectrometry. The new automated sample preparation method increases sample throughput by reducing sample cleanup time to 2 hours for preparing a 96-well plate. The method has excellent sensitivity, specificity, precision (<10%), and accuracy (±15%). We were able to lower the estimated limit of detection (LOD) for COT by 33% and HCT by 73% from our previous LOD. The new LODs for COT and HCT are 0.010 and 0.004 ng/mL, respectively. These lower LODs would enable better detection of SHS in future NHANES surveys.
Immunalysis tapentadol assay reformulation resolves tramadol interference.
Adegoke AO, Jackson AN, La'ulu SL … +4 more , Anderson C, Rudolf JW, Boyd JM, Johnson-Davis KL
J Anal Toxicol · 2025 Nov · PMID 40587090 · Full text
This study evaluated the performance of the Immunalysis Tapentadol 343 Urine Enzyme Immunoassay (EIA) screening kit, focusing on the prevalence of false-positive results due to cross-reactivity with tramadol. Tapentadol...
This study evaluated the performance of the Immunalysis Tapentadol 343 Urine Enzyme Immunoassay (EIA) screening kit, focusing on the prevalence of false-positive results due to cross-reactivity with tramadol. Tapentadol is a dual-action analgesic, modulating μ-opioid receptors and inhibiting norepinephrine reuptake, while tramadol, a structurally related compound, is a weak μ-opioid receptor agonist and norepinephrine/serotonin reuptake inhibitor. Cross-reactivity between these compounds can complicate urine drug screening results for adherence monitoring in chronic pain management. A total of 28 samples initially produced false-positive results for tapentadol BNl using the Immunalysis Tapentadol 343 Urine EIA screening kit. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to confirm the absence of tapentadol. Of the false-positive samples, 61% contained tramadol at concentrations below the manufacturer-reported cross-reactivity threshold of 60 000 ng/mL, indicating assay limitations in specificity. To address this issue, a newly reformulated Immunalysis Tapentadol 343UR Urine HEIA kit was evaluated for tramadol cross-reactivity. Upon retesting the 28 false-positive samples with the reformulated kit, no false positives were detected, with results consistent with LC-MS/MS confirmation. The rate of false-positive tapentadol screens in urine has substantially reduced since the implementation of the new tapentadol kit in routine testing. These findings demonstrate the importance of assay verification to assess cross-reactivity, particularly for structurally related compounds. The reformulated Immunalysis Tapentadol 343UR kit shows improved specificity, reducing false-positive rates and enhancing the accuracy of tapentadol detection in clinical and forensic toxicology applications.
Drug identification in biologicals on clothing, bedding, and other materials.
Labay LM, Midthun KM, Kacinko SL … +1 more , Papsun DM
J Anal Toxicol · 2025 Oct · PMID 40587087 · Publisher ↗
Toxicology testing is an integral component of postmortem, drug-facilitated crime, and driving under the influence investigations. Recommendations pertaining to traditional matrices, sample amounts, and collection contai...
Toxicology testing is an integral component of postmortem, drug-facilitated crime, and driving under the influence investigations. Recommendations pertaining to traditional matrices, sample amounts, and collection container types are well documented in the literature and guidance documents. However, not all cases have traditional toxicological specimens available (e.g. blood with a fluoride additive), and thus require nontraditional toxicology test options. In these cases, a forensic laboratory is contacted to determine if nontraditional objects, such as clothing, bedding, automotive, personal hygiene, or household items, stained with biological material, are suitable for analysis. Comprehensive method validation, as required for routine toxicology tests, is not practical to complete for these items, but this should not deter the toxicology laboratory from taking on this work. Herein, we describe a developed and implemented process for qualitative analysis of biological fluids on/in objects, which ensures the robustness and reliability of reported results. The specific procedures used, which include sample preparation, the incorporation of specialized quality control samples made from the items themselves, analytical acceptance criteria, and reporting considerations are thoroughly detailed. Positive findings from cases were obtained for a variety of drugs, encompassing illicit, prescription, novel psychoactive substances, and over-the-counter medications. Some examples include identification of zolpidem from vomit on clothing; cocaine, cocaine metabolites, levamisole, codeine, acetaminophen, and caffeine in stains on bedding; and diphenhydramine, doxylamine, and dextromethorphan in stains on a mattress pad cover. This methodology is fit-for-purpose and suitable for the toxicological investigation of these unique specimens without any significant limitations. This testing process may be used to identify past drug exposure, associate drug exposure to a particular location or scene, and/or provide insight into an event when a missing person has not been found.
Evaluation of sevoflurane interference with forensic blood ethanol analysis, including sevoflurane stability, and an authentic case.
Perkins C, Rodgers C, Stout P … +1 more , Lee D
J Anal Toxicol · 2025 Oct · PMID 40581784 · Publisher ↗
Sevoflurane, a volatile anesthetic routinely used in clinical settings, was investigated to determine the extent of its interference with in-house forensic blood ethanol analysis. This potential interference could have a...
Sevoflurane, a volatile anesthetic routinely used in clinical settings, was investigated to determine the extent of its interference with in-house forensic blood ethanol analysis. This potential interference could have a significant impact on the analysis and subsequently the interpretation of ethanol in human performance antemortem forensic toxicology casework (e.g. Driving While Under the Influence (DWI) cases). Aqueous samples with ethanol concentrations spanning 0.02-0.40 g/100 mL were fortified with sevoflurane and analyzed using two different dual-column headspace--gas chromatography with flame ionization detection instruments. Sevoflurane was found to elute as an interference peak near ethanol on column 1 (BAC1) and co-elute with ethanol on column 2 (BAC2); the differences were due to the column chemistries. Analyte identification and quantification acceptance criteria monitored included peak-to-valley ratio (resolution) and percent difference between individual column concentrations and the average value of both column concentrations. A 2023 DWI case exhibited potential sevoflurane interference and demonstrated the importance of ethanol reporting acceptance criteria for detecting such interference. In the majority of experiments with sevoflurane and ethanol present in the samples, sevoflurane presence caused failing acceptance criteria to report ethanol results, but if acceptance criteria were met, the ethanol concentration was slightly elevated. An additional sevoflurane stability study showed that the highly volatile sevoflurane could evaporate between analysis and re-analysis of blood samples due to additional tube openings. The decrease of sevoflurane was monitored at each opening of the tube using relative peak areas. HFSC re-analyzes suspected sevoflurane samples, as the additional tube openings could allow sevoflurane to evaporate.
Cannabinoid profiling across toxicology samples in adolescents and young adults by route of administration and in relation to depression symptoms.
Wade NE, Wallace AL, Baca R … +8 more , Andrade G, Happer JP, Courtney KE, Christians U, Sempio C, Klawitter J, Huestis MA, Jacobus J
J Anal Toxicol · 2025 Oct · PMID 40577620 · Full text
Cannabis use is common, with diversity in cannabis products contributing to difficulty in accurately assessing the impact of cannabis use in vulnerable populations such as emerging adults. This study describes and assess...
Cannabis use is common, with diversity in cannabis products contributing to difficulty in accurately assessing the impact of cannabis use in vulnerable populations such as emerging adults. This study describes and assesses concurrence across toxicological matrices (oral fluid, plasma, urine, and hair) and self-reported cannabis use days. Further, it examines whether 11-nor-9-carboxy-tetrahydrocannabinol (THCCOOH, the primary metabolite of Δ9-tetrahydrocannbinol [THC]) concentration or use patterns varies by administration route (smoked flower or vaped concentrate) or predicts depression symptoms. Here, cannabis using (n = 70) and non-using (n = 24) adolescents and young adults (64% female; ages 18-21) were asked to contribute oral fluid, blood, urine, and hair for toxicological testing and self-reported past-90 days of cannabis use, including route of administration. Positive and negative toxicological results by matrix are presented, with sensitivity and specificity calculated. Correlations between THCCOOH concentration across matrices and self-report use were run. Analysis of variance models (ANOVAs) tested whether product type (smoked flower v. vaped concentrate) influenced cannabis use patterns, use to avoid withdrawal, or THCCOOH concentration. Regressions assessed cannabis metrics predicting depression symptoms, controlling for biological sex. All matrices demonstrated excellent specificity (100%), with largely adequate sensitivity (63-74%) except for oral fluid (12%). Self-report and toxicological metrics were significantly correlated (r's = .41-.97), except for avoiding withdrawal. THCCOOH concentration across matrices did not differ by route of administration group; groups also did not differ by self-reported use days or avoiding withdrawal symptoms (p's = .16-.66). Only plasma THCCOOH concentration predicted depression symptoms (beta = 4.43, p < .001). Taken together, toxicological matrices and self-reported cannabis use offer concurrent information in adolescents and young adults who regularly use cannabis. Plasma THCCOOH concentration uniquely predicted self-reported depression symptoms, indicating utility of toxicological cannabinoid concentration predicting clinical outcomes. Given the complexity of measuring cannabis use due to the plethora of available products and rise of new popular cannabinoids, use of toxicological results may offer new insights into clinical outcomes in those who frequently use cannabis.
Cross-reactivity in urine of 53 cannabinoid analogs and metabolites using a carboxylic acid enzyme-linked immunosorbent assay (ELISA) and homogenous enzyme immunoassay (HEIA) kit and immunalysis synthetic cannabinoid HEIA kits.
Yates TL, Poklis JL, Holt AK … +4 more , Fleming JH, Bayard C, Raso SA, Peace MR
J Anal Toxicol · 2025 Oct · PMID 40577619 · Publisher ↗
Advancing knowledge of endocannabinoid receptor agonists and the federal legalization of hemp has created a cannabinoid market of naturally abundant phytocannabinoids to a wide array of semi-synthetic and synthetic canna...
Advancing knowledge of endocannabinoid receptor agonists and the federal legalization of hemp has created a cannabinoid market of naturally abundant phytocannabinoids to a wide array of semi-synthetic and synthetic cannabinoid analogs. Public safety and toxicological concerns exist from lack of regulation, limited pharmacological and metabolomic data, and minimal knowledge of detection ability. Structural similarities of the cannabinoid analogs may allow detection on immunoassays including enzyme-linked immunosorbent assays (ELISA) and homogenous enzyme immunoassays (HEIA), screening platforms in forensic toxicology laboratories for rapid presumptive testing. The cross-reactivity of 27 cannabinoid analogs and 26 commercially available metabolites was evaluated using the Medica EasyRA Enzymatic Immunoassay Analyzer with the Immunalysis Cannabinoids (THC) and Synthetic Cannabinoids 1-3 kits. These analogs were also evaluated using the Dynex DSX Automated ELISA System with the OraSure Technologies Cannabinoids Intercept Microplate EIA. The cannabinoid kits target 11-nor-9-carboxy-Δ9-tetrahydrocannabinol (Δ9-THCCOOH) at a 50 ng/mL cutoff, and the synthetic cannabinoid kits target the N-pentanoic acid metabolite of JWH-018, UR-144, and AB-PINACA at a 10 ng/mL cutoff. Cross-reactivity was evaluated at concentrations of 20, 50, 100, 500, and 1000 ng/mL in urine in triplicate. Absence of cross-reactivity at 1000 ng/mL was considered undetectable. No cross-reactivity was detected on the synthetic cannabinoid kits. Cross-reactivity to Δ9-THCCOOH kits was variable with Δ8-THCCOOH and R-HHCCOOH cross-reacting at the cutoff on the ELISA, with several additional phase I metabolites cross-reacting at 100 ng/mL on both platforms. Analogs lacking the Δ9-THC tricyclic structure and pyran ring cyclization including cannabidiol were undetectable. Alicyclic bond location and alkyl chain length variably affected cross-reactivity, with alkyl lengths 2-4 having increased cross-reactivity comparatively. Compound chirality was also observed to effect instrumental response, with the ELISA having increased cross-reactivity and instrumental response to R-isomers. As knowledge and prevalence of analogs increases, it is crucial to understand the impact on utilized testing platforms.
Tizanidine in postmortem forensic cases.
Friederich LW, Mata DC, Bishop-Freeman SC
J Anal Toxicol · 2025 Oct · PMID 40574679 · Publisher ↗
Alpha-2 adrenergic receptor agonists are established therapeutic medications that are used for conditions such as hypertension, pain disorders, muscle relaxation, spasticity, opioid withdrawal, insomnia, and sedation. Wh...
Alpha-2 adrenergic receptor agonists are established therapeutic medications that are used for conditions such as hypertension, pain disorders, muscle relaxation, spasticity, opioid withdrawal, insomnia, and sedation. While forensic toxicologists may be familiar with more common alpha receptor agonists, tizanidine is a less frequently identified compound, with limited published data available regarding antemortem and postmortem concentrations. Tizanidine therapeutic concentrations found in plasma are reported in the range of 0.0025-0.025 mg/L, although CYP1A2 inhibitors can significantly raise tizanidine levels in the body, thereby increasing the risk of dose-related toxicity. Additionally, imidazoline receptor activity is an underappreciated contributor to the mechanism of action and potential for adverse effects of this drug. Due to its high potency, tizanidine may be missed by forensic laboratories that are not targeting this drug or carefully inspecting untargeted data for its presence. In this study, 18 postmortem cases involving tizanidine are reviewed to improve the understanding of its forensic toxicological profile. These cases have been divided into categories as ruled by the certifying pathologist of "Suicide" involving tizanidine (N = 8, mean 6.2 mg/L and median 0.77 mg/L) and "Accident" involving tizanidine (N = 4, mean 0.86 mg/L and median 0.89 mg/L), and additionally a category of "Incidental" (N = 6, mean 0.35 mg/L and median 0.035 mg/L). Comparison of tizanidine concentrations to those in example cases such as this dataset can assist postmortem forensic toxicologists and pathologists in distinguishing therapeutic postmortem concentrations from toxic/lethal concentrations. However, consideration of scene details and totality of case investigation is essential when determining cause and manner of death.
Incorporation of an environmentally friendly method for the detection of fentanyl and its analogs in oral fluid.
Coulter C, Coulter C, Gonzales J … +1 more , Moore C
J Anal Toxicol · 2025 Nov · PMID 40498581 · Publisher ↗
Oral fluid is considered a favorable matrix for the identification of drug intake mainly because of its simple, observed, non-invasive collection. Fentanyl and fentanyl analog use, misuse, overdose, and deaths are curren...
Oral fluid is considered a favorable matrix for the identification of drug intake mainly because of its simple, observed, non-invasive collection. Fentanyl and fentanyl analog use, misuse, overdose, and deaths are currently occurring at an alarming rate in the USA. The law enforcement community, the Food and Drug Administration (FDA) and the Centers for Disease Control (CDC) are all keenly aware of the urgency in addressing an unmet public health need to identify opioid overdose in individuals as rapidly as possible. As part of a National Institute of Justice grant, the present study was intended to develop and validate an environmentally friendly, rapid, sensitive quantitative method using liquid chromatography coupled to tandem mass spectral detection (LC-MS/MS) for fentanyl and fentanyl analogs in oral fluid collected using the nform rapid test device. Oral fluid samples were subjected to liquid-liquid extraction incorporating bio-renewable solvents where possible, reducing the environmental footprint of the assay. A buffer/salt free mobile phase was employed consisting of 0.1% formic acid in water (95%): 0.1% formic acid in methanol (5%) at a flow rate of 0.8 mL/min; the run time was 4.5 minutes, again reducing environmental impact in terms of salt and solvent usage. The method included fentanyl, 4-anilino-N-phenethylpiperidine; (4-ANPP; desproprionyl fentanyl), acetyl fentanyl, carfentanil, p-fluorofentanyl, valeryl fentanyl, p-fluorobutyrylfentanyl, furanyl fentanyl and benzoyl fentanyl as well as xylazine, which is often detected with fentanyl. The method was validated according to ANSI/ASB 036 (2019) Standard Practices for Method Validation in Forensic Toxicology.
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