Drug Testing and Analysis最新文献

Hormonal growth promotants (HGPs) are a class of pharmaceutical agents commonly administered to cattle in the United States to improve growth rates of the animal, alter behavior, or to improve the desired characteristics of retail cuts of meat. There is a concern that low residual concentrations of HGPs may remain in tissue after slaughter, and consumption of tissues containing these compounds may increase the risk of adverse health outcomes, including cancer. Sensitive and selective methods are necessary to assess exposure of HGPs by populations that consume meat products from animals that may have been administered HGPs. A liquid chromatography-tandem mass spectrometry method was developed and validated to detect the low-level presence of HGPs including estradiol, testosterone, estradiol benzoate, melengestrol, melengestrol acetate, progesterone, testosterone propionate, trenbolone, trenbolone acetate, and α-zearalanol in retail cuts of meat following a liquid-liquid extraction using a high pH solution with 30-50× less mass of meat required as compared to similar approaches. Good chromatographic performance and sensitivity was achieved utilizing ammonium fluoride as a mobile phase additive without the need for derivatization. Validation parameters including accuracy, precision, recovery, matrix effects, limits of detection, limits of quantitation, linear range, and stability were determined. The limits of detection ranged from 0.1 to 1.0 ng/g, depending on the compound, with adequate accuracy and precision without the need for extensive sample preparation approaches.

Erythropoietin receptor agonist (ERA) testing for antidoping can be achieved in both urine and blood samples. Recent published work showed the comparability between the two matrices and focused on detectability in microvolumetric capillary serum samples collected using the Tasso+ device. Currently, in the antidoping field, blood samples are required to be shipped under cold, temperature-controlled conditions. However, due to the suggested greater stability of EPO in blood compared to urine, it is believed that blood samples should be viable for ERA analysis if shipped under the ambient, not cold temperature-controlled conditions to which urine samples are subjected. In this collaborative study with the Ultimate Fighting Championship, microvolumetric capillary serum samples were collected in the field and shipped under ambient conditions to the laboratory for ERA analysis. Resulting data showed that endogenous EPO was detectable in 100% of these samples, showing no loss in detectability despite shipping under non-controlled conditions. Further, ERA analyses were conducted in the laboratory on additional in-house collected samples and post-EPO administration samples subjected to various storage and shipping conditions, also showing reliable endogenous and recombinant EPO detectability in all samples except those experiencing extreme temperature (50°C) conditions. Taken together, these data highlight the stability of EPO in blood samples and show that ERA blood samples can be collected in the field and shipped without costly temperature-controlled shipping methods and without a loss in detectability.

The phosphodiesterase 4 (PDE4) inhibitors constitute a relatively modern class of medications that are known for inducing bronchodilation and exhibiting anti-inflammatory properties within the body. Due to these properties, there is concern regarding their potential misuse as performance-enhancing substances in competitive sports. This study delves into the metabolic conversion of roflumilast in thoroughbred horses following oral administration and in vitro experimentation using equine liver microsomes and Cunninghamella elegans. High-performance liquid chromatography coupled with a Q Exactive Orbitrap mass spectrometer (HPLC-HRMS) was employed for analysis. The investigation identified 10 metabolites of roflumilast, including six phase I and four phase II metabolites from in vivo studies, and 11 metabolites from in vitro studies, consisting of eight phase I and three phase II metabolites. The identified biotransformation products encompassed processes such as hydroxylation, chlorine substitution, methylation, N-oxide formation, and even the dissociation of methylenecyclopropane and difluoromethane. Furthermore, the study identified three glucuronic acid and one sulfonic acid conjugated phase II metabolites of the investigated drug candidate. The aforementioned findings contribute to the detection and comprehension of the unauthorized utilization of roflumilast in equestrian sports.

We developed a method for comprehensive urine drug screening by applying dilute-and-shoot extraction and vacuum-insulated probe-heated electrospray ionization with ultra-high performance liquid chromatography high-resolution quadrupole time-of-flight mass spectrometry (DS-UHPLC-VIP-HESI-QTOFMS). The method involved five-fold post-hydrolysis dilution of urine samples and chromatography on a C18 UHPLC column prior to QTOFMS analysis. The recently introduced VIP-HESI ion source was chosen due to its enhanced ionization efficiency and compatibility with UHPLC-QTOFMS. Extensive data was acquired in positive ion mode with a low collision energy (7 eV) and an elevated collision energy (30 eV), using the broadband collision-induced dissociation data acquisition scan mode that continuously generated high-resolution and accurate mass for parent and fragment qualifier ions, and parent ion isotopic patterns. Compound identification was performed against an in-house database with 1263 compound entries, using an automated post-run reverse target database search with preset identification criteria. Method validation with 56 different drugs showed acceptable results for the limit of identification (median 5 ng/mL), matrix effects (70-130%), repeatability of retention times (< 1%), mass accuracy (< 1 mDa), as well as for specificity and stability. As compared with an established UHPLC-QTOFMS method relying on solid-phase extraction and conventional electrospray ionization, DS-UHPLC-VIP-HESI-QTOFMS produced comparable results from authentic clinical urine samples for most drugs, but showed clearly improved detectability for pregabalin, gabapentin, and ritalinic acid. We anticipate that the new method will be a step forward for laboratories performing routine urine drug screening due to its fast turnaround time, reduced manual workload, cost efficiency, and broad substance coverage.

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).