Drug Testing and Analysis最新文献

The increasing misuse of opioids in Europe is an alarming trend, leading to severe social and health consequences. Heroin, a highly potent and addictive opioid, remains the main contributor to the health burden associated with opioid use in the region. Illicit drug characterization and profiling offer valuable insights into the complexity of heroin seizures, assisting law enforcement agencies and forensic experts in gathering evidence for legal proceedings. This study provides a comprehensive overview of the composition of heroin seizures and assesses the feasibility of an electrochemical fingerprint approach for the detection of heroin and its associated components. In the initial phase, the primary focus was on developing an electrochemical sensor optimized for heroin detection. The sensor's performance was validated using street samples provided by Sciensano, a Belgian health institute, ensuring its accuracy and reliability in identifying heroin. Once the capabilities of the sensor were demonstrated, the discrimination of alkaloids and cutting agents in seized samples was integrated into a customized software script. Subsequently, an extensive validation process was conducted using a new dataset of heroin seizures from the Belgian National Institute for Criminalistics and Criminology. The follow-up verification confirmed the sensor's effectiveness in detecting heroin, cutting agents, and alkaloids, highlighting its potential as a valuable tool for drug profiling. This portable, user-friendly device with automatic readout could become essential for forensic experts, law enforcement, and harm reduction efforts in addressing the opioid crisis.

Athletes are increasingly using probiotic supplementation to support their overall health, and it can be particularly beneficial for female athletes in managing recurrent urinary tract infections and bacterial vaginosis. One route of probiotic administration for females is vaginal application, which enables direct modulation of the microbiota. While probiotics are widely recognised for their health benefits, their potential impact on urinary steroidal markers monitored in the Steroidal Module of the Athlete Biological Passport remains unexplored. Given the biological overlap between vaginal and urinary microbiomes, bacteria from vaginal probiotics could transfer into urine samples, potentially altering steroid profiles through microbial enzymatic activity. This study investigates whether vaginal probiotic use, specifically Lactobacillus reuteri and plantaraium, could influence urinary steroid markers relevant to the steroidal passport. In vitro and in vivo approaches were employed to evaluate the potential effects of contamination and variability on key steroidal markers. Analyses of in vitro and in vivo experiments suggest that vaginal probiotics do not substantially affect urinary steroid markers monitored in the Athlete Biological Passport. However, some variations were observed that merit further investigation. These findings contribute to a better understanding of how vaginal probiotics might interfere with doping control results, emphasising the need for further research to ensure accurate interpretation of urine steroidal profiles in the female athlete.

For the past couple of years, black market products have appeared and were confirmed to contain genetic products coding for human erythropoietin (EPO). While being prohibited by the World Anti-Doping Agency (WADA), they could be used to produce endogenously more EPO hormone and hence increase performance. In a previous work, we demonstrated the potential of 20-μL dried blood spots (DBS) to detect the presence of EPO transgene in human blood down to 250 copies (12,500 copies/mL), despite lower sensitivity (30-fold) than in 1-mL fresh blood. As the use of DBS as a collection matrix for antidoping is going to expand in the near future, our aim was to develop and validate a new protocol to improve the sensitivity of gene doping detection from DBS. Three DBS devices were evaluated: polymeric Tasso-M20 (TASSO Inc.) and Mitra (Neoteryx), and cellulosic Protein Saver 903 (Whatman). The best results were achieved with polymeric DBS, and a full validation was performed for the detection of the EPO transgene using Tasso M-20 DBS; 1500 copies/mL were detected in 50% of cases and robust detection was obtained at 5000 copies/mL (100 copies transgene in 20-μL DBS) with the four spots of the Tasso device tested over several weeks. The results confirm that polymeric DBS can be used as an alternative to fresh blood for gene doping detection with high sensitivity simplifying also potential reanalysis in the future.

An innovative software tool for the rapid and efficient simulation of the metabolism of new psychoactive substances (NPS) was developed, based on the open-source project mzmine, and applied. NPS are compounds designed to mimic the psychotropic effects of established illicit drugs while circumventing drug legislation. These compounds are developed solely regarding their desired effects, thus possibly leading to harmful side effects including the formation of toxic metabolites. Analytical reference standards, needed to carry out metabolic studies, are not immediately available because emerging NPS are primarily discovered subsequent to drug confiscations. Using these confiscated substances in traditional metabolic in vivo or in vitro studies is often not possible due to the substances being impure or being a part of a mixture of different NPS. Therefore, a software tool was developed to streamline the evaluation of data acquired by the online combination of electrochemistry and mass spectrometry for the simulation of NPS metabolism. Using this tool, it is possible to generate mass voltammograms directly from mass spectrometric raw data. Combining this newly implemented tool with existing filtering algorithms in mzmine, we simulated the metabolism of the synthetic cannabinoid receptor agonist (SCRA) methyl 3,3-dimethyl-2-[1-(pent-4-en-1-yl)-1H-indazole-3-carboxamido] butanoate (MDMB-4en-PINACA) from a mixed solution of different NPS. Fragmentation data indicated that one of the transformation products found for MDMB-4en-PINACA is likely of a quinoid structure. The potential formation of this possibly highly reactive quinoid metabolite could be a first hint for possible causes of adverse side effects frequently reported after the recreational use of MDMB-4en-PINACA and related SCRAs.

New psychoactive substances (NPS), especially synthetic cannabinoid receptor agonists (SCRA), are increasingly smuggled into prisons via infused mail. Consumption of those substances by inmates in prisons is associated with increased aggression, violence, and organized crime. Onsite detection of infused mail often is challenging. Because the infused papers do not show any visible stains or olfactory changes, physical inspection is often insufficient. The applicability of further conventional on-site detection methods like immunoassays and sniffer dogs is severely limited. Because of the rapidly changing supply of already circulating and newly emerging NPS, it is impractical to impossible to keep up with the development of immunoassays or the training of sniffer dogs. Hence, confiscated mail samples are routinely tested by either liquid chromatography or gas chromatography coupled to mass spectrometry (MS), which is costly and time-consuming. In this study, recent advancements in the hyphenation of laser ablation (LA) and molecular MS were investigated regarding the possible application for the rapid and easy detection of NPS in prison mail. Utilizing an in-house developed LA-MS hyphenation based on atmospheric pressure chemical ionization (APCI), 31 mail samples confiscated in German prisons were analyzed. It was possible to correctly identify 27 samples containing SCRAs. For these positive samples, it was also possible to detect the specific compounds each paper was infused with. The use of LA-APCI-MS has simplified sample preparation and reduced analysis time per sample to 1 min.

Hypoxia-inducible factor prolyl hydroxylase inhibitors (HIF-PHIs) represent a novel class of therapeutic substances that increase erythropoiesis. Due to their performance-enhancing effects and potential risk of abuse, these agents were added to the World Anti-Doping Agency (WADA) Prohibited List in 2011. Enarodustat is a novel HIF-PHI and has been approved for clinical use in China in 2023. This study primarily aimed to characterize its major urinary metabolites for antidoping purposes. A single oral dose of 40-mg enarodustat was administered to a volunteer. Urine samples were collected over 28 days and processed using solid-phase extraction (SPE). Analytical methods included liquid chromatography-high resolution mass spectrometry (LC-HRMS) under both positive and negative electrospray ionization conditions, complemented by in vitro metabolism studies using human liver microsomes (HLMs) for the characterization and identification of metabolites. A total of eight metabolites were detected, including Phase I products such as parent compound (PC) isomer (M1), monohydroxylation (M2), dihydroxylation (M3), and dehydrogenation (M4) metabolites, as well as Phase II conjugates involving methylation (M5), glycosylation (M6), glucuronidation (M7), and monohydroxylation-sulfation (M8) in vivo. Among these, M3-M5 are novel metabolites. In addition, compared with other metabolites, PC and M2 exhibited longer detection windows, suggesting they are valuable biomarkers for doping control purposes. The study elucidates enarodustat's metabolic pathways and provides a foundation for developing sensitive detection methods. Future work should focus on synthesizing reference materials to identify metabolite structures.

Phenibut is a new psychoactive substance (NPS) first synthesized in Russia in 1963 as a derivative of gamma-aminobutyric acid. Originally developed for therapeutic use, it has gained popularity for nonmedical purposes, including recreational and cognitive enhancement. In Brazil, phenibut is uncontrolled and easily purchased online. This study used wastewater-based epidemiology (WBE) to investigate phenibut use patterns in two northeastern Brazilian cities. Composite daily wastewater samples were collected from two treatment plants (WWTPs), Recife (WWTP_A) and Olinda (WWTP_B), during two periods in 2023: Carnival and a reference week. Samples underwent solid-phase extraction (SPE) and analysis by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Phenibut concentrations were converted to population-normalized mass loads (PNMLs, mg/day/1000 inhabitants). The highest phenibut levels and PNMLs (up to 4.06 mg/day/1000 inhabitants) occurred during Carnival at WWTP_A, located in a major tourist area, suggesting recreational use. During the reference week, PNMLs ranged from detection limits to 2.29 mg/day/1000 inhabitants on weekdays, indicating possible functional or cognitive enhancement use. These findings reveal two distinct use patterns: recreational peaks during Carnival weekends and possible functional use on weekdays outside festive periods. This is the first evidence of phenibut detection in Brazilian wastewater and its temporal use patterns. The results highlight WBE's value in monitoring NPS trends and suggest recreational use predominates during large events. This underscores the need for public health attention and regulatory monitoring of uncontrolled substances with abuse potential.

Cannabis sativa is one of the oldest and most versatile plants with many facets ranging from intoxicant to medicine. Legalisation of medicinal cannabis leads to an increasing complexity of specific forensic questions to distinguish between recreational and medicinal use, for example, in context with participation in road traffic. Hence, there is a recent interest in finding objective markers that enable the differentiability of cannabis flowers. Terpenes, volatile hydrocarbons with a modular construction principle of isoprene subunits, are currently suggested as a second substance class alongside phytocannabinoids for the classification of cannabis material. A headspace full evaporation technique gas chromatography mass spectrometry (HS-FET-GC/MS) methodology was successfully validated according to forensic guidelines for the analysis of 45 terpenes in cannabis flowers including 16 monoterpenes, 16 monoterpenoids, 7 sesquiterpenes and 6 sesquiterpenoids. FET-sampling was developed in detail experimentally, revealing evidence of thermal instability of higher-boiling terpenes. Validation included selectivity, linearity of calibration (ranges 10–2000 μg/g), analytical limits (at least 6 μg/g), accuracy (bias) as well as intraday and interday precision. The use of a retention time index mixture as an internal standard and measurement in SIM-scan mode also allows for the qualitative identification of further terpenes present in cannabis. Application to a set of cannabis strains with similar Δ⁹-THC content demonstrated differences and similarities in their terpene profiles.

Using a targeted metabolic investigation approach, a new, previously undescribed metabolite, which is a pyrrole derivative of LGD-4033, has been detected and coded as M8. This metabolite can be detected in postadministration human urine samples up to 6 days after administration. It has also been detected in post-administration samples, mimicking supplement contamination, after repeated 10 μg doses detectable for ≥ 120 h after administration. Given M8's structural similarity to LGD-4033, its androgen receptor (AR) agonist/antagonist properties were studied using in silico molecular docking and functional in vitro AR transactivation assays in the PC3(AR)₂ cell model, alongside other selected LGD-4033 metabolites. The results indicate that M8 can act as a potent AR antagonist, whereas M2c was reconfirmed as a potent AR agonist. Therefore, we propose the inclusion of M2c in ITP doping control methods, as it could be used as an LGD-4033 alternative and may be introduced into the black market. Additionally, the detection of M8, which is an early-stage excreted metabolite, is valuable for estimating sample collection time relative to LGD-4033 intake. When combined with the evaluation of other long-term metabolites like M5b, M5a, M2c, and M2d, M8 detection can aid in distinguishing adverse analytical findings, associated abuse through regular dosing, from unintentional doping caused by certain contamination scenarios or abuse through microdosing.