Drug Testing and Analysis最新文献

In this study, the performance of the new MAIIA EPO purification kit 7D3 was evaluated for human urine samples to confirm compliance with the WADA criteria of the WADA technical document 2024 (TD2024EPO). The kit was validated for selectivity, reliability at minimum required performance levels (MRPLs), limit of detection (LOD), carryover and recovery. A total of 81 urine samples were tested for selectivity, 29 samples for reliability at MRPL, LOD and 16 samples for recovery. The kit fulfils the criteria and the requirements of the upcoming WADA TD2024EPO and presents a valid solution for the purification of ERAs from human urine for doping analyses.

β-hydroxy β-methyl butyric acid (HMB), either as the free acid or in the form of its calcium salt, is a component of several dietary supplements marketed to enhance sports performance, due to its role in protein synthesis. In this pilot study, we investigated the urinary excretion profile of HMB, an endogenous metabolite of the essential amino acid leucine. The endogenous reference ranges of HMB, in human urine samples, were monitored by collecting samples from 20 volunteers. Data obtained were compared with those from controlled excretion studies, following the intake of a 3-g oral dose of HMB. Urine samples were analyzed through a targeted procedure employing a conventional GC-MS system operating in SIM mode. Our results show that the excreted urinary concentration of HMB significantly exceeds the range of endogenous concentrations for at least 24 h, making possible to detect its exogenous origin.

In forensic toxicology, it has been debated if hair testing allows an estimation of the intensity of cocaine use-an assumption that may have risen because self-reports in a forensic setting are of uncertain validity per se. We therefore investigated the relationship between self-reported cocaine use and cocaine hair concentrations (including its main metabolites benzoylecgonine and norcocaine) in chronic cocaine users voluntary participating in psychiatric study settings. Additionally, we tested whether hair testing can distinguish between individuals with and without a diagnosis of cocaine dependency. Cocaine users (N = 195) from three independent experimental studies reported their average powder cocaine consumption in g/week over the last 3-4 months in an interview and provided a 3- to 4-cm hair sample assayed with liquid chromatography tandem-mass spectrometry. Moreover, study participants were assessed with the Structured Clinical Interview (SCID-IV) for psychiatric diagnoses. Using linear regression models, we found a robust correlation between cocaine_total (sum of cocaine and metabolites) hair concentration and self-reported cocaine use in g/week (r_cocainetotal = 0.47, p < 0.001), indicating that 1000 pg/mg cocaine_total corresponded to a use of 0.80 g/week (confidence interval [95%]: 0.56-1.07 g/week). In logistic regression models, cocaine_total hair concentration predicted cocaine dependency with a sensitivity of 0.79 and a specificity of 0.65 (threshold 0.5), suggesting its acceptable capacity to distinguish dependent from non-dependent cocaine users. The findings may have significant implications for forensic and clinical practices, encouraging the use of hair analysis as a potential tool for monitoring cocaine use and dependence.

In the United States, melatonin products are widely available as dietary supplements. During the past few decades, melatonin products have gained popularity primarily as a sleep aid, with a variety of product forms available for different age groups of consumers. Recent reports have highlighted a rise in melatonin ingestion among children reported to poison control centers. The increased use of melatonin-containing products, the diversity in product forms, and reported label discrepancies has emphasized the need for additional research to better understand the marketplace. This work aims to measure melatonin content in products sold as dietary supplements and marketed for children, evaluate method performance across different product categories, and determine if product form has an impact on melatonin stability. One hundred ten (110) dietary supplement products labeled to contain melatonin and marketed towards children were purchased and analyzed using a targeted LC-MS/MS method validated for the qualitative determination of serotonin and quantification of melatonin, N¹-acetyl-N²-formyl-5-methoxykynuramine (AFMK), and N¹-acetyl-5-methoxykynuramine (AMK). Melatonin was identified in 108 of 110 products (98%) with a median concentration of 1.2 mg/g (range: 0.017-130 mg/g) or 1.7 mg/serving (range: 0.042-50 mg/serving). Further, in the tested products, melatonin content ranged from 0% to 667% of the label declaration. This study provides evidence to inform safety assessments and investigate potential factors that may influence reported concentrations, such as product stability and matrix influences.

Vamorolone, a potential alternative to conventional glucocorticoids, shows significant promise in sports medicine due to its reduced side effects and superior pharmacodynamic properties. This study aims to investigate the metabolic characteristics of this novel synthetic cyclodextrin-steroid anti-inflammatory drug and elucidate the metabolic pathways in human liver microsomes (HLMs) in vitro, thereby providing a scientific basis for assessing its potential risks for athletes. All compounds are detected by liquid chromatography-high resolution mass spectrometry (LC-HRMS) and metabolite identification was performed using Compound Discoverer 3.3 software. In the HLMs model, 12 metabolites of vamorolone are successfully identified, including 10 phase I metabolites and 2 phase II metabolites. Among these, the reduction metabolite M1 exhibited the highest peak area, indicating it as one of the primary metabolic pathways. The dehydrogenated compound M2 had the second highest peak area, further elucidating the metabolic characteristics of vamorolone. This study systematically identifies the metabolite structures of vamorolone in HLMs and provide crucial data for the pharmacokinetics and biomarker research of this drug. The findings not only enhance the understanding of its metabolic mechanisms but also offer a scientific basis for evaluating its safety and efficacy in sports medicine. Meanwhile, these discoveries can contribute to better regulation and control of Vamorolone's use in competitive sports, ensuring fairness in competitions.

Ethyl glucuronide (EtG) in hair is a reliable biomarker of alcohol consumption habits. Due to its small concentration incorporated into hair, analytical methods sensitive enough to reliably quantify EtG in this matrix are required. Sample preparation is critical in hair analysis, especially for EtG, for which extraction efficiency and matrix effect can strongly influence the results; furthermore, miniaturized methods are sought, to reduce solvent use and times of sample preparation. A micro extraction by packed sorbent (MEPS) procedure coupled to a high-performance liquid chromatography-tandem mass spectrometry method was developed and validated for quantitation of EtG in human hair samples. Fifty milligrams of hair samples were cut into snippets and extracted in water. The cleanup of the extract was carried out by using a MEPS syringe packed with anion exchange sorbent (SAX); all parameters for conditioning, washing, loading and eluting steps were optimized and the eluted aqueous volume was directly injected in the LC-MS/MS system operating in the negative ionization mode. The method was fully validated assessing LOD, LOQ, calibration curve, repeatability, accuracy, matrix effect and carryover. The method was subsequently applied to QCs and authentic hair samples. The developed MEPS method is quick and effective, with low solvent purchase and discard costs, allowing the differentiation between social drinkers and chronic excessive alcohol consumers, according to the cut-offs established by the Society of Hair Testing (SoHT).

The amino-terminal propeptide of type III procollagen (P-III-NP) is used with IGF-I to detect the illicit use of growth hormone and to monitor growth hormone therapy. However, the only currently available assays for P-III-NP are immunoassays, which are not well harmonized. In addition, other fragments of type III procollagen may better evaluate collagen turnover. We aimed to develop a high-throughput assay using immunoaffinity enrichment coupled to ultra-high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) to quantify peptides belonging to three different regions of type III procollagen in human serum simultaneously. To facilitate higher throughput, we transferred the assay from microcentrifuge tubes to a 96-well plate format with partially automated pipetting. The method was linear (Pearson's R ≥ 0.994) over an estimated concentration range of 1.35-13.3 nM, 0.04-2.28 nM, and 0.26-5.1 nM for each surrogate peptide of P-III-NP, collagen degradation products, and the carboxyl-terminal propeptide, respectively. Intra-day and inter-day imprecision were both < 13.6%, and the results of robustness testing were also encouraging. The method was successfully applied to capillary blood samples obtained using Tasso+ microsampling devices. Modest correlation of P-III-NP concentration was observed between our new method and a WADA-approved immunoassay (N = 40, Pearson's R = 0.789) with a significant bias of -87.8%. Our method simultaneously quantifies four peptides belonging to three regions of type III procollagen in human serum. High bias between assays highlights the need for common higher-order calibrators or reference materials to help improve the comparability of results across laboratories.

Blood collection is an important facet of anti-doping testing, forming the basis of the hematological module of the athlete biological passport (ABP). Presently, whole blood is collected via venepucture under standardized conditions, in accordance with the World Anti-doping Agency's International Standard for Testing and Investigations (ISTI). Advances in capillary whole blood collection technology now afford the ability to collect microvolumetric capillary whole blood from the upper arm (or other suitable vascular location such as the abdomen) that is "needle-free" and virtually painless. Previous work has demonstrated the suitability and feasibility of capillary whole blood compared with venous using the Tasso+ device. Recently, other microcapillary devices have received FDA approval increasing the options available to the anti-doping community. The present study extends previous work, by collecting microliter capillary whole blood samples via two different FDA approved devices (RedDrop and the Tasso+ device) to compare with venous blood collected simultaneously. Ten participants provided three matched blood samples (2× capillary and 1× venous) in accordance with WADA ABP guidelines for blood sample collection, for a total of 30 samples. Capillary samples from both devices showed excellent laboratory agreement with venous blood for all CBC parameters, with the exception of platelets. Excellent laboratory agreement was also observed between the two microcapillary collection devices. Irrespective of the device, microcapillary blood collection provides a valid alternative to venous blood collection for ABP purposes.

Detecting the use of bisphosphonates (BPs) in equine athletes is of interest to regulators and laboratories due to the threat to welfare issues for the potential to provide analgesic effects and manipulating bone structure. The detection of BPs in biological matrices is challenging due to erratic biological elimination and inconsistent analytical recoveries. Therefore, complementary approaches are needed to provide evidence of their misuse in racehorses. BPs have two sub-classes: nitrogenous and non-nitrogenous. This study investigated plasma elimination following administration of one example from each sub-class, together with changes in endogenous eicosanoid and corticosteroids. Zoledronic acid (ZA) and tiludronic acid (TA) were administered by IV infusion to 8 thoroughbred horses with an 11-month washout period between each administration. Sample preparation for quantification of BPs by liquid chromatography-tandem mass spectrometry (LC-MS/MS) utilised a two-step solid phase extraction (SPE) consisting of polymeric reversed-phase followed by weak anion exchange prior to derivatisation using trimethyl orthoacetate. Endogenous biomarkers were analysed after protein precipitation and SPE with polymeric reversed-phase prior to liquid chromatography-high resolution mass spectrometry (LC-HRMS) using data independent acquisition. The LC-MS/MS analysis showed ZA was undetectable after 8 h post-administration while TA was detected up to the final collection point of 28 days post-administration. The LC-HRMS analysis utilised targeted (i.e., prior inclusion list of compounds) approaches to monitor level changes of eicosanoid and corticosteroid biomarkers. Putative biomarkers were identified and now subject to validation for translation into routine sample analysis for improved retrospectivity to detecting BP misuse in equine plasma.

After the Scottish Prison Service (SPS) introduced mail photocopying procedures in December 2021, a shift in smuggling methods was observed for synthetic cannabinoid receptor agonists (SCRAs) and other new psychoactive substances (NPS) from drug-infused papers back to traditional sample matrices (e.g., tablets and powders), although new matrices also emerged. This study reports on waxy- or putty-like materials as a novel drug preparation for SCRAs and other drugs seized from UK prisons. In 2023, 22 of these new preparations were seized from Scottish prisons, with eight found in sealed vape pods. The materials were positive for SCRAs, phytocannabinoids, novel benzodiazepines, and/or gabapentinoids. Additionally, 11 preparations were seized from an English prison, all containing the SCRAs MDMB-4en-PINACA and MDMB-INACA. MDMB-INACA was pharmacologically characterized using in vitro CB₁ and CB₂ bioassays, revealing moderate efficacy but low potency at CB₁. Furthermore, the in vitro CB₁ bioassay was also used to evaluate the CB₁ activating potential of extracts from eight seized samples. Six of these showed high CB₁ activity, whereas the samples lacking SCRAs or containing only MDMB-INACA showed no or only weak CB₁ activity, respectively. Lastly, applying the bioassay as an activity-based "untargeted" screening method effectively identified the presence of SCRAs in one waxy preparation, which was initially not detected by gas chromatography-mass spectrometry (GC-MS). This underscores the effectiveness of the bioassay for evaluating these new waxy- or putty-like materials for the presence of SCRAs.