Journal of analytical toxicology最新文献

Benzimidazole opioids (nitazenes) are novel synthetic opioid receptor agonists that over the last few years have emerged on recreational drug markets, and their abuse has become a concern worldwide. In particular, limited documentation of their pharmacology and toxicology, along with their unpredictable presence in counterfeit medicines mistaken for established brands, pose significant challenges. Herein, we present a case of fatal intoxication with the nitazene etonitazepyne, after intake of tablets appearing like, and thus mistaken as, oxycodone. The assertion of etonitazepyne's implication in the case was delayed by several months, due to lack of information about and access to seized tablets. Eventually, an analytical method using liquid chromatography coupled to a Waters Xevo TQ-S tandem quadrupole mass spectrometer (LC-MSMS) was developed and documented. Using this method, etonitazepyne was confirmed in the tablets and quantified at a concentration of 0.32 ng/mL in both femoral blood and vitreous fluid sampled at autopsy of the deceased. The femoral blood concentration is in the lower range compared to previously published etonitazepyne-related deaths. This case illustrates the challenges with detecting nitazenes and the imminent health risk counterfeit products poses, even for experienced drug users.

Analytical toxicology is a discipline of forensic toxicology which applies analytical techniques for the determination of drugs of abuse in biological and non-biological 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.

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.

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.

Trazodone, a medicine registered for human, is a serotonin agonist-antagonist. At low dose, the drug is sedative due to its antagonist properties. At high dose, it is an agonist with anxiolytic and antidepressant actions. Trazodone can be administered to the horse to reduce anxiety. However, according to the antidoping rules for horses, the presence of trazodone in blood or urine is considered as a violation, which will produce a suspension of both the athlete and the horse as the drug is listed banned on the International Federation of Horseracing Authorities prohibited substances list. As a hair test can provide more evidence or supplementary information to an adverse analytical finding or to document drug exposure, our forensic laboratory received two specimens with a request for trazodone identification. After mane collection, trazodone was analysed by a new LC-MS/MS method involving pH 9.5 borate buffer overnight incubation of 20 mg of specimen in presence of clozapine-d4 used as internal standard, followed by solvents extraction. Linearity was verified from 1 to 100 pg/mg (R2 = 0.9967). Limit of detection of the method was 0.1 pg/mg. Trazodone was measured at 0.4 pg/mg in the mane of a horse suspended after an antidoping violation. In a case of hidden administration, trazodone was identified at 9 and 24 pg/mg in two consecutive mane hair segments. Although no controlled study allows interpretation, particularly about the frequency of exposure and the dose that entered in the body, this is the first evidence that trazodone can be incorporated in the mane of horses.

Synthetic cathinones (SCs) were confirmed as the second most prevalent class of New Psychoactive Substances (NPS) in 2024, underscoring their widespread availability and use. Notably, the SCs seizure and related intoxication cases increased within the European Union, involving mostly 3-chloromethcathinone, 3-methylmethcathinone, 2-methylmethcathinone, and N-ethylnorpentedrone. Italy has observed a distinct trend, with methylendioxy pyrrolidinohexanophenone (MDPHP) emerging as a significant concern. This technical note aims to provide a comparative analysis of the most recent data concerning the emergence and the spread of MDPHP in Italy. Data from the National Early Warning System on Drugs indicate that the total amount of seized MDPHP increased from 2021 to 2024. In 2023, MDPHP-related intoxication cases peaked at 47, considering either alone or in combination with other drugs, followed by a slight decrease in 2024. In Italy, MDPHP seems to dominate the Italian black market of SCs and with two fatalities reported. This increased demand for MDPHP raises health concerns, indicating the necessity to enhance drug-related harm reduction services and improve a multidisciplinary network that provides data for understanding the complex phenomenon of NPS.

4-Phosporyloxy-N, N-dimethyltryptamine (psilocybin) is a psychedelic tryptamine found in certain mushroom species that has shown efficacy in the treatment of various psychiatric disorders. In conjunction with the renewed interest in therapeutic effects of psychedelics, there has been an increase in psilocybin-like designer tryptamines appearing in non-medical drug markets. The present study aimed to develop and validate a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for detecting and quantifying 4-position ring-substituted tryptamines and their 4-hydroxy metabolites in plasma. Specifically, we investigated 4-phosphoryloxy-N, N-dimethyltryptamine (psilocybin), 4-acetoxy-N, N-dimethyltryptamine (psilacetin), 4-propionoxy-N, N-dimethyltryptamine (4-Pro-DMT) and their shared metabolite 4-hydroxy-N, N-dimethyltryptamine (psilocin), along with 4-methyl carbonato-N, N-di-n-propyltryptamine (4-MeCO3-DPT) and its metabolite 4-hydroxy-N, N-di-n-propyltryptamine (4-HO-DPT). Mass spectrometry analysis employed electrospray ionization (ESI) in positive mode, with two multiple reaction monitoring (MRM) transitions per analyte. Plasma samples were acidified with ascorbic acid, followed by protein precipitation with acetonitrile. Linearity was achieved across a concentration range of 0.5-100 ng/mL for all analytes, except psilocybin, which displayed linearity from 5 to 100 ng/mL. Validation results demonstrated acceptable bias (±20%) and imprecision (<20%) for all analytes. Matrix effects, evaluated in 10 samples (CV <18.3%), indicated minimal interference, although ion enhancement was observed for psilocin (31.9%) and psilocybin (45.7%). Extraction efficiency across all tryptamines was approximately 50%. The assay method was used to quantitate plasma samples from male rats treated with 1.0 mg/kg s.c. of the prodrug psilacetin, and collected before and 5, 30, 60, 120 and 240 min after injection. No psilacetin was detected, and psilocin concentrations ranged from non-detected up to 32.7 ng/mL. Overall, we successfully developed a sensitive and specific method for the detection and quantification of six tryptamines in plasma, providing a robust tool for future research and clinical applications.

Postmortem alcohol production by microorganisms has been known to increase blood alcohol concentration, potentially leading to erroneous interpretation. Here, we present a peculiar case where postmortem toxicology detected a high ethanol concentration only in the stomach content. An 87-year-old bedridden woman was taken to senior day care facility, where the nurse attempted to take her vitals but noticed she was not breathing. The facility called for an ambulance, but she was pronounced dead soon after arriving at the hospital. She was found to be severely dehydrated, and her blood tests indicated poor health. Autopsy was performed 2 days after death, and postmortem alcohol analysis detected a high ethanol concentration only in the stomach content. Our aim was to investigate the causative agent of ethanol production; microbiological analysis identified the yeast species Candida glabrata as the responsible microorganism.

A drug-related fatality involving 3,4-methylenedioxy-α-pyrrolidinohexanophenone (MDPHP) is here reported. Belonging to the class of synthetic cathinones (SCs), MDPHP is a 3,4-methylenedioxy-derived designer (MDDs) drug with a pyrrolidine moiety and an alkyl portion with six carbon atoms. Other MDD pyrrolidine derivatives belong to the alkyl homologous series (C3-C5) and are known as 3,4-methylenedioxy-α-pyrrolidinopropiophenone (MDPPP), 3,4-methylenedioxy-α-pyrrolidinobutyrophenone (MDPBP) and 3,4-methylenedioxypyrovalerone (MDPV). MDDs are psychostimulant drugs of abuse that primarily act on monoamine transporters; little is known about their off-target liability. Recently, MDPHP has gained attention due to increasing seizures and involvement in human intoxications, but currently there is a lack of data about its pharmaco-toxicological effects. In the case reported here, a 58-year-old man with a history of MDPV addiction was found dead in a waterway. While no evidence of natural disease or trauma was found to account for the death, toxicological analysis revealed the presence of MDPHP in addition to MDPPP, MDPV, MDPBP, clonazepam, and citalopram. Since no standards of MDPPP and MDPBP were available at the time of the analysis, LC-QTOF analysis of the drugs and their metabolites were performed. The following concentrations of MDPHP were reported: 350 ng/mL in femoral blood (FB), 110 ng/mL in cardiac blood (CB), 1900 ng/mL in urine, 3000 ng/mL in bile, 490 ng/g in kidney, 80 ng/g in liver, 480 ng/g in lung, 98 ng/g in brain, 700 ng/mL in gastric content and 8 ng/mg in pubic hair. Other MDDs concentrations in biological fluids and tissue were significantly lower than MDPHP suggesting their presence as synthetic impurities. Finally, to better understand the binding properties of the abovementioned MDDs to several documented transporters and receptors, an in silico evaluation was performed. The medical examiner reported that the cause of death was an acute multidrug intoxication by MDPHP and clonazepam in presence of MDPPP, MDPV, MDPBP and citalopram.

A streamlined liquid chromatography quadrupole time-of-flight mass spectrometry method utilizing protein precipitation and filtration extraction was developed to achieve rapid and reliable screening and confirmation for blood and urine matrices. This method targets 946 drugs and metabolites across 35 drug classes via sequential window acquisition of all theoretical mass spectra with variable customized windows to enhance spectral clarity, and was validated per established guidelines to ensure high accuracy and reproducibility. Combined with complementary in-house methods, this approach meets and exceeds the testing requirements outlined in ANSI/ASB standards and recommendations for postmortem, drug-facilitated crime, and Tier I and II driving under the influence of drug analyses. The method demonstrated efficient and sensitive performance, achieving limits of detection as low as 0.1 ng/mL. It accurately identified expected detections across 67 proficiency test samples and 224 authentic case samples, with high accuracy and reliability in the detection of both traditional drugs and novel psychoactive substances. The method employs an in-house built library and incorporates in-batch standards analyzed alongside case samples to ensure contemporaneous identification criteria, making it suitable for confirmation and reporting purposes. By expanding the analytical capabilities to include a vast range of analytes, this method improves the likelihood of identifying substances that may otherwise go undetected and reduces the need for multiple separate tests, thereby enhancing the overall effectiveness of toxicological investigations.