Drug Testing and Analysis最新文献

Gas chromatography–combustion–isotope ratio mass spectrometry (GC-C-IRMS) is the method of choice to detect the abuse of synthetic forms of naturally occurring steroids for doping control purposes. GC-C-IRMS relies on a multistep sample cleanup to ensure each target analyte peak is chromatographically pure before combustion. To achieve that, liquid–liquid or solid phase extraction (SPE) is commonly used in combination with preparative liquid chromatography (LC).

In this work, a procedure for isolation, purification, and analysis of endogenous steroids by GC-C-IRMS was developed and validated. The key elements were successive application of strong cation and strong anion exchange SPE with enzymatic hydrolysis in between to strip the ionic species from urine and decrease matrix complexity prior to LC cleanup; preparative two-dimensional LC, where only the testosterone fraction required secondary purification (40 min total run time per sample); and derivatization of selected fractions with formic acid to yield formate esters, followed by GC-C-IRMS analysis. Formylation afforded excellent separation between 5α- and 5β-androstanediols and simplified the detection of the endogenous reference compound pregnanetriol by converting it to a volatile artifact, tentatively identified as 3α,20-diformoxy-17-methyl-18-nor-5β,17α-pregn-13-ene.

The overall method performance benefited from the customization of the GC-C-IRMS combustion interface, which improved robustness and facilitated the transfer of sample components into the hot zone of the oxidation reactor, minimizing peak tailing. The former was achieved by keeping the oxygen flow through the reactor at all times, obviating the need for periodic oxidation, and the latter—by implementing a direct capillary-in-capillary connection of the chromatographic column to the oxidation reactor.

This paper describes the detection of nandrolone decanoate and its metabolites in mane hair collected from horses that have been treated with nandrolone decanoate (Deca-Durabolin). The intramuscular administration study of nandrolone decanoate in three Thoroughbred castrated horses (each received 800 mg weekly for 3 consecutive weeks) was previously conducted to investigate its metabolism and detection time in plasma and urine for doping control purposes. In this work, segmental analysis of the post-administration hair has revealed that (i) nandrolone decanoate and its metabolites, nandrolone and 4-estrene-3,17-dione, could be detected in horse mane after intramuscular administration, and (ii) the spreading of these substances along the entire hair strand was observed, which suggested the involvement of sweat or sebum incorporation.

Although incineration is currently the primary method for the disposal of seized illicit drugs, alternative methods for the disposal of illicit drugs may be necessary to provide safer and more accessible alternatives. Chemical oxidation processes have been identified as a promising alternative method to degrade illicit drugs. Using commercially available reagents and established industry processes, chemical degradation holds potential as an alternative drug disposal technique. This study investigated the oxidants ozone, sodium hypochlorite, trichloroisocyanuric acid, hydrogen peroxide, OXONE, sodium percarbonate, and peracetic acid for their potential to degrade illicit amphetamine-type stimulants using β-phenethylamine (PEA) as an exploratory analog. Oxidants and conditions that showed the highest degradation efficiency with PEA were applied to methamphetamine, amphetamine, 3,4-methylenedioxymethamphetamine (MDMA), and 3,4-methylenedioxyamphetamine (MDA). Transformation products were identified using gas chromatography–mass spectrometry, and degradation was quantified using a fit-for-purpose method via liquid-chromatography quadrupole time-of-flight mass spectrometry.

Of the oxidants explored, ozone performed the best, leading to high degradation efficiencies of methamphetamine (95%), amphetamine (86%), MDMA (100%), and MDA (100%) after 72 h of exposure. Sodium hypochlorite was also highly effective for the degradation of methamphetamine and amphetamine, while trichloroisocyanuric acid was particularly effective for MDMA and MDA. All the major transformation products of degradation were tentatively identified, with only one of 10 listed as a controlled, scheduled, or restricted substance. This research demonstrates how chemical degradation can provide a novel alternative to incineration for the destruction of amphetamine-type stimulants, providing a sustainable, long-term, and accessible method of illicit drug disposal.

Epristeride, a novel noncompetitive inhibitor of Type II 5α-reductase, has emerged as a potential therapeutic alternative for benign prostatic hyperplasia (BPH). Given that other 5α-reductase inhibitors, such as finasteride and dutasteride, are already monitored for their potential impact on doping control, comprehensive metabolic studies of epristeride are crucial for antidoping. This study investigates the metabolic pathways and metabolites of epristeride using in vitro microsome models, offering preliminary insights into the pharmacokinetics of this drug. Metabolite profiling was performed using liquid chromatography–high resolution mass spectrometry (LC-HRMS), with data acquisition facilitated by Xcalibur 4.2 software and metabolite identification facilitated by Compound Discoverer 3.3. By employing network pharmacology, the potential targets of epristeride are predicted. The binding energy is calculated using AutoDock Vina software to predict its impact on steroid metabolism. The study proposed three primary metabolites of epristeride: two Phase I oxidation products (M1 and M2) and one Phase II glucuronidation product (M3). Pathway analysis revealed that among the five CYP450 isoforms examined, CYP3A4 played a dominant role. The docking results tentatively elucidated five key target proteins (ESR1, CYP19A1, STAT3, AKR1C3, and CYP17A1) with low binding energies, indicating stable interactions. Notably, Phase I metabolites (M1 and M2) showed significant binding potential with these targets, whereas the Phase II metabolite (M3) exhibited lower binding stability. These findings provide a detailed understanding of epristeride's metabolic pathways and its potential biological impacts, offering valuable insights for monitoring its presence as a confounding factor in doping control.

Beta₂-adrenergic agonists are widely used for bronchial relief in respiratory conditions such as asthma and exercise-induced bronchoconstriction. However, this drug class has also been shown to have muscle anabolic properties. Kelch-Like Family Member 41 (KLHL41) is a sarcomeric protein implicated in muscle remodeling and hypertrophy. In this study, we examined the effects of oral clenbuterol, therapeutic inhaled formoterol, and resistance training on KLHL41 protein abundance in human skeletal muscle. KLHL41 levels were measured by immunoblotting in vastus lateralis muscle biopsies from healthy adults following 2 weeks of oral clenbuterol administration, 6 weeks of inhaled formoterol at therapeutic doses, or 8 weeks of resistance training. We also assessed sex differences and the effects of acute versus prolonged interventions. Prolonged oral clenbuterol administration significantly increased KLHL41 abundance compared to placebo (p < 0.001), with a magnitude similar to that observed after resistance training (p < 0.01), whereas therapeutic inhaled formoterol had no effect on KLHL41 levels. Neither acute clenbuterol administration nor a single resistance training session altered KLHL41 abundance, and no sex differences were observed in baseline KLHL41 levels. These findings indicate that beta₂-adrenergic stimulation via oral clenbuterol, but not therapeutic inhalation of formoterol, promotes sarcomeric remodeling through KLHL41-related pathways similar to those activated by resistance training. The distinct effects of these agents on KLHL41 support current anti-doping regulations prohibiting clenbuterol use and highlight KLHL41 as a potential molecular marker of skeletal muscle adaptation to hypertrophic stimuli.