Search

The Brazilian National Metrology Institute produced a suite of certified reference materials (CRMs) intended as internal standards (ISs) for quantitative nuclear magnetic resonance (qNMR). Being a ratio primary method, the use of qNMR in organic chemistry has already crossed the borders of research laboratories, despite the cost of instrumentation. The International Bureau of Weights and Measures (BIPM) proposed eight potential qNMR ISs. Four candidate materials were selected for their solubility in various solvents and distinct chemical shifts, making them suitable for qNMR analysis of diverse analytes. The certification process compared orthogonal primary methods such as mass balance, qNMR, freezing-point depression, and coulometry, to ensure independent value assignment. Different approaches were compared to assess batch homogeneity and stability. While directly comparing the main compound's chromatographic area proved to be a quick and fit-for-purpose approach, the determination of individual impurities provided lower uncertainties but required more laborious work. CRM batches of maleic acid, dimethyl sulfone, potassium hydrogen phthalate, and dimethyl terephthalate were delivered with over 999.8 mg g -1 purity and uncertainty in the range of 0.6 to 3 mg g -1 (k = 2). The literature shows certification procedures for qNMR ISs whose traceability chain is exclusively based on qNMR measurements. As opposed to that, the methodology presented here provides robust certified values assigned by methods independent of qNMR, in accordance with BIPM recommendations and less prone to qNMR biases. The CRMs developed in this work have already been used for SI-traceable purity evaluation of compounds such as drugs and pesticides, by laboratories in Brazil and abroad., Competing Interests: Declarations. Conflict of interest: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH, DE part of Springer Nature.)

Introduction: Data Fusion-based Discovery (DAFdiscovery) is a pipeline designed to help users combine mass spectrometry (MS), nuclear magnetic resonance (NMR), and bioactivity data in a notebook-based application to accelerate annotation and discovery of bioactive compounds. It applies Statistical Total Correlation Spectroscopy (STOCSY) and Statistical HeteroSpectroscopy (SHY) calculation in their data using an easy-to-follow Jupyter Notebook., Method: Different case studies are presented for benchmarking, and the resultant outputs are shown to aid natural products identification and discovery. The goal is to encourage users to acquire MS and NMR data from their samples (in replicated samples and fractions when available) and to explore their variance to highlight MS features, NMR peaks, and bioactivity that might be correlated to accelerated bioactive compound discovery or for annotation-identification studies., Results: Different applications were demonstrated using data from different research groups, and it was shown that DAFdiscovery reproduced their findings using a more straightforward method., Conclusion: DAFdiscovery has proven to be a simple-to-use method for different situations where data from different sources are required to be analyzed together., (© 2022 John Wiley & Sons Ltd.)

Certified reference materials (CRM) of amphetamine derivatives were produced through a simple, rapid and efficient synthesis in both batch and continuous-flow conditions, accompanied by the development of a comprehensive certification protocol for this class of substances. Our chemistry enabled the synthesis of MDA, MDMA, PMA and PMMA in two steps from safrole and estragole with overall yields of 38-61 % in 48 hours under batch conditions and 61-65 % in 65 minutes under continuous-flow conditions, followed by the development of a certification protocol for these materials through identity checking, homogeneity, stability, and characterization studies. Furthermore, as result of this work, a very pure CRM of MDA.HCl with 99.1±1.4 g/100 g of certified characterization value was produced. Considering the importance of supplying amphetamine calibrants for public security efforts in Forensic Chemistry, the potential therapeutical applications, and responding to the rising demand for the synthesis of CRM, this work presents a pioneering approach for the production of amphetamine and related compounds., (© 2023 Wiley-VCH GmbH.)

Comparability of measurement results and their metrological traceability to the International System of Units (SI) are fundamental tools to ensure reliable decisions in the social sphere, commerce, and science. The use of appropriate references in analytical chemistry, such as certified reference materials (CRMs) of high purity substances, is one of the required procedures to obtain traceable measurements. When commercial standards with non-certified purity values are used, traceability must be achieved by determining the purity of the standard using a potential primary reference measurement procedure or other appropriate methods. Quantitative nuclear magnetic resonance (qNMR) is a technique with the potential to be used in primary measurement procedures. This work presents the determination of purity by 1 H qNMR for nitrofuran metabolites 3-amino-2-oxazolidinone (AOZ), 3-amino-5-morpholinomethyl-2-oxazolidinone (AMOZ), and 1-aminohydantoin (AHD). Furthermore, a recent qNMR method developed by our group to improve the quantitative performance of measurements using 13 C nucleus was used to determine the purity of semicarbazide (SEM) nitrofuran metabolite. Purity values obtained by qNMR for AOZ, AMOZ, and AHD standards were compared to values obtained by the mass balance approach using a suite of analytical methods: Karl Fischer (KF) coulometric titration and thermogravimetry (TG) for the determination of water and residual solvents, gas and liquid chromatography for the determination of impurities structurally related to the metabolites. The results obtained by qNMR and mass balance were consistent.Graphical abstract.

The estimation of limits of detection (LOD) for solely qualitative methods in analytical chemistry may prove challenging because all the approaches with which chemists are familiar require some type of numeric data input. The best model to describe the binary response in these methods (detected/not detected) is a logistic model; however, these models are not easily handled by most of the laboratories and generally demand expensive statistical software packages. In this work, the advantages of applying this approach are discussed and its implementation using commercial spreadsheet software is demonstrated. A free online application based on the R environment using shinyapps was developed and its application was validated and discussed with a dataset of 57 different target compounds analyzed in urine according to the requirements of the World Anti-Doping Agency (WADA). This tool allows free, extremely quick, and easy determinations of LOD in qualitative analyses as well as the determination of the probabilities of detection in any given concentration., (© 2022 John Wiley & Sons Ltd.)

Introduction: In this era of 'omics' technology in natural products studies, the complementary aspects of mass spectrometry (MS)- and nuclear magnetic resonance (NMR)-based techniques must be taken into consideration. The advantages of using both analytical platforms are reflected in a higher confidence of results especially when using replicated samples where correlation approaches can be used to statistically link results from MS to NMR., Objectives: Demonstrate the use of Statistical Total Correlation (STOCSY) for linking results from MS and NMR data to reach higher confidence in compound identification., Methodology: Essential oil samples of Melaleuca alternifolia and M. rhaphiophylla (Myrtaceae) were used as test objects. Aliquots of 10 samples were collected for GC-MS and NMR data acquisition [proton ( 1 H)-NMR, and carbon-13 ( 13 C)-NMR as well as two-dimensional (2D) heteronuclear single quantum correlation (HSQC), heteronuclear multiple-bond correlation (HMBC), and HSQC-total correlated spectroscopy (TOCSY) NMR]. The processed data was imported to Matlab where STOCSY was applied., Results: STOCSY calculations led to the confirmation of the four main constituents of the sample-set. The identification of each was accomplished using; MS spectra, retention time comparison, 13 C-NMR data, and scalar correlations of the 2D NMR spectra., Conclusion: This study provides a pipeline for high confidence in compound identification using a set of essential oils samples as test objects for demonstration., (© 2022 John Wiley & Sons, Ltd.)

Unlabelled: The growing consumption of nutritional supplements might represent a problem, given the concern about the quality of these supplements. One of the most used supplements is whey protein (WP); because of its popularity, it has been a target of adulteration with substitute products, such as cheaper proteins with lower biological value. To investigate this type of adulteration, this study used shotgun proteomics analyses by MS(E) (multiplexed, low- and high-collision energy, data-independent acquisition) of WP-based supplements. Seventeen WP-based supplement samples were evaluated. Chicken, maize, rice, potato, soybean, and wheat proteins were considered as probable sources of bovine whey adulteration. Collectively, 523 proteins were identified across all 16 samples and replicates, with 94% of peptides inside a normal distribution within 10ppm of maximum error. In 10 of the 16 samples analyzed, only proteins from bovine whey could be detected, while in the other samples several other protein sources were detected in high concentrations, especially soybean, wheat, and rice. These results point out a probable adulteration and/or sample contamination during manufacturing that could only be detected using this proteomic approach., Significance: The present work shows how shotgun proteomics can be used to provide reliable answers in quality control matters, especially focusing on Whey Protein nutritional supplements which are a very popular subject in food and nutrition. In order to achieve an appropriate methodology, careful evaluation was performed applying extremely rigorous quality criteria, established for the proteomic analysis. These criteria and the methodological approach used in this work might serve as a guide for other authors seeking to use proteomics in quality control., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Verapamil and naproxen Parallel Artificial Membrane Permeability Assay (PAMPA) permeability was studied using lipids not yet reported for this model in order to facilitate the quantification of drug permeability. These lipids are 1,2-distearoyl-sn-glycero-3-phosphatidylcholine (DSPC), 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) and an equimolar mixture of DMPC/DSPC, both in the absence and in the presence of 33.3 mol% of cholesterol. PAMPA drug permeability using the lipids mentioned above was compared with lecithin-PC. The results show that verapamil permeability depends on the kind of lipid used, in the order DMPC > DMPC/DSPC > DSPC. The permeability of the drugs was between 1.3 and 3.5-times larger than those obtained in lecithin-PC for all the concentrations of the drug used. Naproxen shows similar permeability than verapamil; however, the permeability increased with respect to lecithin-PC only when DMPC and DMPC/DSPC were used. This behavior could be explained by a difference between the drug net charge at pH 7.4. On the other hand, in the presence of cholesterol, verapamil permeability increases in all lipid systems; however, the relative verapamil permeability respect to lecithin-PC did not show any significant increase. This result is likely due to the promoting effect of cholesterol, which is not able to compensate for the large increase in verapamil permeability observed in lecithin-PC. With respect to naproxen, its permeability value and relative permeability respect lecithin-PC not always increased in the presence of cholesterol. This result is probably attributed to the negative charge of naproxen rather than its molecular weight. The lipid systems studied have an advantage in drug permeability quantification, which is mainly related to the charge of the molecule and not to its molecular weight or to cholesterol used as an absorption promoter.

Quantitative nuclear magnetic resonance measurements have become more popular over the last decade. The introduction of new methods and experimental parameters has been of fundamental importance in the development of new applications. Amongst these new developments is the introduction of electronic referencing for quantifications. The use of electronic referencing eliminates errors in the analyses as a result of weighting of internal standards as well as undesired problems as a result of the solubility of the standards in the analyte solution and chemical interactions between the analyte and the internal standard. In this work, we have studied the quantification of a very important analyte in a food matrix, benzoic acid in orange juice, as a model to the validation and measurement uncertainty estimation of electronic referencing using (1)H NMR in food analyses. The referencing method applied was the pulse length-based concentration measurement. Method was validated and showed good results for the precision and accuracy parameters evaluated. A certified reference material and a reference material candidate were analyzed, and extremely good results were obtained. Reported relative expanded uncertainties are in the 1.07-1.39% range that can be considered an extremely good performance for the analysis of a food complex matrix. Measurement uncertainty was evaluated by two different approaches, and the pulse calibrations for the samples and for the reference have been shown to account for approximately 80% of the total uncertainty of the measurement., (Copyright © 2014 John Wiley & Sons, Ltd.)

Salbutamol is commonly used in asthma treatment, being considered a short-effect bronchodilator. This drug poses special interest in certain fields of chemical analysis, such as food, clinical and doping analyses, in which it needs to be analyzed with quantitative precision and accuracy. Salbutamol, however, is known to degrade under certain conditions and this is critical if quantitative results must be generated. The present work aimed to investigate salbutamol extraction from urine samples, to determine whether salbutamol is unstable in other solvents as well as in urine samples, to elucidate the structures of the possible degradation products and to validate an analytical method using the extraction procedure evaluated. Stability investigations were performed in urine at different pH values, in methanol and acetone at different temperatures. Semi-preparative liquid chromatography was performed for the isolation of degradation products, and gas chromatography coupled to mass spectrometry as well as nuclear magnetic resonance were used for identification. Three unreported methylation products were detected in methanolic solutions and had their structures elucidated. Urine samples showed a reduction in salbutamol concentration of up to 25.8% after 5 weeks. These results show that special care must be taken regarding salbutamol quantitative analyses, since degradation either in standard solutions or in urine could lead to incorrect values., (Copyright © 2013 John Wiley & Sons, Ltd.)

Certified reference materials (CRMs) are essential tools to guarantee the metrological traceability of measurement results to the International System of Units (SI), which means the accuracy and comparability of results over time and space. In the pharmaceutical area, only a few CRMs are available and the use of (non-certified) reference materials is a much more common practice. In this paper, the studies on a new candidate CRM of sodium diclofenac based on the ISO Guides 34:2009 and 35:2005 are described. The project steps included characterization, homogeneity test, stability studies, and uncertainties estimation. In the characterization, the mass fractions of organic, inorganic, and volatile impurities were determined, and the results were cross-checked by independent reference methods or interlaboratorial study. The API mass fraction was calculated by mass balance and cross-checked by quantitative proton nuclear magnetic resonance (¹H qNMR). The paper also presents a Monte Carlo simulation to estimate the measurement uncertainty as an approach to validate the GUM results in ¹H qNMR. The homogeneity between batch units was verified, and the candidate CRM stability under transport and storage conditions was evaluated in short- and long-term stability studies. The CRM certified property value and corresponding expanded uncertainty, obtained from the combined standard uncertainty multiplied by the coverage factor (k=2), for a confidence level of 95%, was (999.76+0.10) mg g⁻¹., (Copyright © 2012 Elsevier B.V. All rights reserved.)

The use of "nutritional supplements" containing unapproved substances has become a regular practice in amateur and professional athletes. This represents a dangerous habit for their health once no data about toxicological or pharmacological effects of these supplements are available. Most of them are freely commercialized online and any person can buy them without medical surveillance. Usually, the steroids intentionally added to the "nutritional supplements" are testosterone analogues with some structural modifications. In this study, the analyzed product was bought online and a new anabolic steroid known as methylstenbolone (2,17α-dimethyl-17β-hydroxy-5α-androst-1-en-3-one) was detected, as described on label. Generally, anabolic steroids are extensively metabolized, thus in-depth knowledge of their metabolism is mandatory for doping control purposes. For this reason, a human excretion study was carried out with four volunteers after a single oral dose to determine the urinary metabolites of the steroid. Urine samples were submitted to enzymatic hydrolysis of glucuconjugated metabolites followed by liquid-liquid extraction and analysis of the trimethylsilyl derivatives by gas chromatography coupled to tandem mass spectrometry. Mass spectrometric data allowed the proposal of two plausible metabolites: 2,17α-dimethyl-16ξ,17β-dihydroxy-5α-androst-1-en-3-one (S1), 2,17α-dimethyl-3α,16ξ,17β-trihydroxy-5α-androst-1-ene (S2). Their electron impact mass spectra are compatible with 16-hydroxylated steroids O-TMS derivatives presenting diagnostic ions such as m/z 231 and m/z 218. These metabolites were detectable after one week post administration while unchanged methylstenbolone was only detectable in a brief period of 45 h., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Exemestane is an aromatase enzyme complex inhibitor. Its metabolism in humans is not fully described and there is only one known metabolite: 17β-hydroxyexemestane. In this work, excretion studies were performed with four volunteers aiming at the detection of new exemestane metabolites in human urine by gas chromatography coupled to mass spectrometry (GC-MS) after enzymatic hydrolysis and liquid-liquid extraction. Urine samples collected from four volunteers were analyzed separately. The targets of the study were mainly the 6-exomethylene oxidized metabolites. Two unreported metabolites were identified in both free and glucuconjugated urine fractions from all four volunteers, both of them were the result of the 6-exomethylene moiety oxidation: 6ξ-hydroxy-6ξ-hydroxymethylandrosta-1,4-diene-3,17-dione (metabolite 1) and 6ξ-hydroxyandrosta-1,4-diene-3,17-dione (metabolite 2). Furthermore, only in glucoconjugated fractions from all volunteers, one metabolite arising from the A-ring reduction was identified as well, 3ξ-hydroxy-5ξ-androst-1-ene-6-methylene-17-one (metabolite 3). The molecular formulae of all these metabolites were ascertained by the determination of exact masses using gas chromatography coupled to high resolution mass spectrometry (GC-HRMS). Moreover, all metabolites were confirmed using an alternative derivatization with methoxyamine and MSTFA/TMS-imidazole., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Trimethylsilylation of anabolic agents and their metabolites is frequently achieved by using the derivatization mixture N-methyl-N-(trimethylsilyl)trifluoroacetamide (MSTFA)/NH(4)I/2-mercaptoethanol. Nevertheless, artifacts were formed when this mixture was employed in the monitoring of exemestane and its main metabolite 17β-hydroxyexemestane prior to GC-MS analysis. These artifacts were identified as the N-methyltrifluoroacetamide (MTFA) and trimethylsiloxyethylmercapto products of the respective trimethylsilyl (TMS) derivatives. Furthermore, artifact formation was evaluated taking the structure (1,4-diene-3-keto-6-exomethylene) of the compounds into account. Although these artifacts are relevant for investigations regarding the derivatization process and may be of interest in many fields, they are detrimental to cope with the requirements of the World Anti-Doping Agency (WADA) in terms of the limits of detection (LODs) required. To overcome this issue, a method using an alternative derivatization was proposed: formation of methyloxime-TMS derivatives through double derivatization using O-methylhydroxylamine/pyridine and MSTFA/TMS imidazole after enzymatic hydrolysis and liquid-liquid extraction. Samples from an excretion study after administration of exemestane to healthy volunteers were analyzed by the proposed method and detection of both exemestane and its main metabolite was possible. This method showed excellent results for both analytes meeting the LODs required for antiestrogenic agents (50  ng/mL) established by WADA. The method was validated for the main metabolite, it was robust and cost-effective for qualitative and quantitative purposes, with LOD and LOQ of 10  ng/mL and 25  ng/mL, respectively., (2010 John Wiley & Sons, Ltd.)

Certified reference materials (CRMs) contribute to the traceability of measurement results to recognized anchor points, ideally to the International System of Units (SI), and consequently, to their comparability and reliability. In the area of veterinary drugs' residues in foods of animal origin, the important role of CRMs must be highlighted, given the problems caused by trade restrictions in some countries. The results of studies concerning the certification of an incurred matrix certified reference material (IMCRM) for the mass fraction of nitrofuran metabolites in chicken muscles are presented in this work. A batch of IMCRM candidates was produced from chicken muscle samples containing the metabolites 3-amino-2-oxazolidinone (AOZ), 3-amino-5-morpholinomethyl-2-oxazolidinone (AMOZ), semicarbazide (SEM), and 1-aminohydantoin (AHD). All the statistical approaches for the homogeneity and stability assessments were based on the principles contained within ISO Guide 35 (current ISO 33405:2024). All the quantitative analyses of the nitrofuran metabolites were performed by means of high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). The properties studied were sufficiently homogeneous and stable at -20 °C for 360 days (the defined storage conditions). Under transport conditions, the AOZ metabolite showed instability at 50 °C. The residual moisture content was also monitored and showed instability at 20 °C and 50 °C. The characterization was performed via an exact matching calibration approach and the assigned values with their respective expanded uncertainties (k = 2) were 5.40 ± 1.2 ng g -1 , 5.49 ± 0.85 ng g -1 , 5.08 ± 1.2 ng g -1 , and 6.01 ± 0.85 ng g -1 for the metabolites AOZ, AMOZ, AHD, and SEM, respectively, in the lyophilized chicken muscle. These values ranged from 1.3 to 1.5 ng g -1 on a wet basis. The expiration date for the CRM was estimated based on the stated limits for future values calculated from the confidence interval of the stability study's linear regression curve., Competing Interests: Declarations. Ethics approval: This study used samples of animal origin. An ethics approval was not necessary because the entire process, including breeding, feeding, and slaughter, was carried out following the same procedures used for animals intended for food production. Furthermore, a veterinary doctor monitored the entire process. Conflict of interest: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH, DE part of Springer Nature.)

Nuclear magnetic resonance (NMR) has fast emerged as a quantitative platform for it offered numerous advantages over chromatographic methods in analyzing purity and determining content of organic compounds. However, the mechanism of parameter settings has not been further investigated causing the improper or incorrect use in the widespread qNMR applications. This study aims to construct a quantitative 1H NMR methodology based on the experiment optimization and simulation and explain the impact of parameter settings on accuracy. We successfully applied the proposed 1H qNMR method with the internal standard method to assess the purity of ofloxacin, a commonly used antibacterial drug. The study showed that parameter settings, including flip angle, relaxation delay, transmitter frequency offset, scan number and data point, had different effects on the quantification errors (0.5 % to 8.9 %). The method validation proved that the 1H qNMR methodology delivered high accuracy with the measurement uncertainty (0.60 %). The results demonstrated that the proposed 1H qNMR method provided clear guidance on the practical implementation of accurate quantification of low molecular mass compounds (< 500 g/mol). [ABSTRACT FROM AUTHOR]

Cobalt was included on the World Anti‐Doping Agency Prohibited List in 2015 due to its effect on stimulus of erythropoiesis via stabilization of hypoxia‐inducible factor. Although it has proven benefits for performance enhancement, the unavailability of inductively coupled plasma–mass spectrometry on routine of the accredited laboratories is a factor that reduces its applicability in anti‐doping analysis. Therefore, an analytical method for quantification of urinary cobalt as its diethyldithiocarbamate complex by liquid chromatography coupled with high‐resolution tandem mass spectrometry was developed and validated. Palladium was proposed as internal standard and rhodium as a complexation control. A microwave‐assisted acid digestion of the urine samples was essential, not only to eliminate the matrix effect but mainly to avoid the non‐specific bond of cobalt to endogenous molecules. A linear method was obtained over the studied range from a negative urine control to a spiked concentration of 25 ng/mL, with an estimated limit of quantification of 2.5 ng/mL, and an adequate combined standard uncertainty of 11.4%. Considering that all reagents are commercially available, the proposed strategy is feasible to be included on routine sample preparation. Monitoring urinary cobalt concentrations globally opens the perspective to support the anti‐doping system to define a suitable threshold value and to understand its potential misuse by athletes seeking for performance improvement. [ABSTRACT FROM AUTHOR]

Higenamine is prohibited in sports as a β2‐agonist by the World Anti‐Doping Agency. As a key component of a great variety of plants, including the Annonaceae family, one aim of this research project was to evaluate whether the ingestion of Annona fruit could lead to higenamine adverse analytical findings. Single‐dose administration studies including three Annona species (i.e., Annona muricata, Annona cherimola, and Annona squamosa) were conducted, leading to higenamine findings below the established minimum reporting level (MRL) of 10 ng/mL in urine. In consideration of cmax values (7.8 ng/mL) observed for higenamine up to 24 h, a multidose administration study was also conducted, indicating cumulative effects, which can increase the risk of exceeding the applicable MRL doping after Annona fruit ingestion. In this study, however, the MRL was not exceeded at any time point. Further, the major urinary excretion of higenamine in its sulfo‐conjugated form was corroborated, its stability in urine was assessed, and in the absence of reference material, higenamine sulfo‐conjugates were synthesized and comprehensively characterized, suggesting the predominant presence of higenamine 7‐sulfate. In addition, the option to include complementary biomarkers of diet‐related higenamine intake into routine doping controls was investigated. A characteristic urinary pattern attributed to isococlaurine, reticuline, and a yet not fully characterized bismethylated higenamine glucuronide was observed after Annona ingestion but not after supplement use, providing a promising dataset of urinary biomarkers, which supports the discrimination between different sources of urinary higenamine detected in sports drug testing programs. [ABSTRACT FROM AUTHOR]

Proficiency testing is the use of interlaboratory comparisons to assess the ability of a laboratory to perform a particular test or measurement competently and demonstrate the reliability of the results generated. To achieve this goal, the international standard ISO 13528:2022 presents four performance statistics and one of them is the z score. Some researchers have proposed an alternative approach for proficiency testing scheme interpretation based on an analysis of variance method. This short communication presents residual analysis to choose an appropriate analysis of variance model for proficiency testing interpretation when the assigned value is obtained by consensus value among participants. The analysis of variance approach was compared with z score to assess the performance of interlaboratory comparison for determining the amount of NOx present in vehicle emissions. Preliminary results indicate that the conclusions obtained by both methodologies differ for one of the participating laboratories. This occurs due to violation of the homoscedasticity assumption which is not considered when using the z score. On the other hand, residual analysis can provide such information and feasible generalized least squares can deal with heteroscedasticity. [ABSTRACT FROM AUTHOR]

Proficiency testing schemes by interlaboratory comparisons are used to determine the performance of individual laboratories for specific tests or measurements. The international standard ISO 13528:2015 provides a description of statistical methods used for achieving this goal; one of these methods is the z score. The standard allows each participant to choose a measurement method and this can lead to inhomogeneity between participant variance known as heteroscedasticity. The ISO 13528:2015 standard does not mention heteroscedasticity in its statistical procedures. This paper describes a new approach, based on residuals analysis, to assess the performance of an interlaboratory comparison for determining the presence of benzoic acid in orange juice. The results indicate that the conclusions (using z scores) and the proposed approach are different for some laboratories. This occurs due to violation of the homoscedasticity assumption. The z score procedure does not consider this assumption violation while residual analysis can provide such information. Feasible generalized least squares allow one to deal with non-homoscedasticity. [ABSTRACT FROM AUTHOR]

The presence of bio-macromolecules as major ingredients is a primary factor in marketing many biologically derived macromolecular supplements. Workflows for analyzing these supplements for quality assurance, adulteration, and other supply-chain difficulties must include a qualitative assessment of small-molecule and macromolecular components; however, no such integrated protocol has been reported for these bio-macromolecular supplements. Twenty whey protein supplements were analyzed using an integrated workflow to identify protein content, protein adulteration, inorganic elemental content, and macromolecular and small-molecule profiles. Orthogonal analytical methods were employed, including NMR profiling, LC-DAD-QToF analysis of small-molecule components, ICP-MS analysis of inorganic elements, determination of total protein content by a Bradford assay, SDS-PAGE protein profiling, and bottom-up shotgun proteomic analysis using LC-MS-MS. All 20 supplements showed a reduced protein content compared to the claimed content but no evidence of adulteration with protein from an unclaimed source. Many supplements included unlabeled small-molecule additives (but nontoxic) and significant deviations in metal content, highlighting the importance of both macromolecular and small-molecule analysis in the comprehensive profiling of macromolecular supplements. An orthogonal, integrated workflow allowed the detection of crucial product characteristics that would have remained unidentified using traditional workflows involving either analysis of small-molecule nutritional supplements or protein analysis. [ABSTRACT FROM AUTHOR]

Development of diagnostic testing capability has advanced with unprecedented pace in response to the COVID-19 pandemic. An undesirable effect of such speed is a lack of standardization, often leading to unreliable test results. To assist the research community surmount this challenge, the National Research Council Canada has prepared a SARS-CoV-2 spike protein reference material, SMT1-1, as a buffered solution. Value assignment was achieved by amino acid analysis (AAA) by double isotope dilution liquid chromatography–tandem mass spectrometry (LC-ID-MS/MS) following acid hydrolysis of the protein, in combination with ultraviolet–visible spectrophotometry (UV–Vis) based on tryptophan and tyrosine absorbance at 280 nm. Homogeneity of the material was established through spectrophotometric absorbance readings at 280 nm. Transportation and long-term storage stabilities were assessed by monitoring relative changes in oligomeric state by size-exclusion liquid chromatography (LC-SEC) with UV detection. The molar concentration of the spike protein in SMT1-1 was 5.68 ± 0.22 µmol L−1 (k = 2, 95% CI), with the native trimeric form accounting for ~ 94% of the relative abundance. Reference mass concentration and mass fraction values were calculated using the protein molecular weight and density of the SMT1-1 solution. The spike protein is highly glycosylated which leads to analyte ambiguity when reporting the more commonly used mass concentration. After glycoprotein molar mass determination by LC-SEC with multi-angle light scattering detection, we thus reported mass concentration values for both the protein-only portion and intact glycoprotein as 0.813 ± 0.030 and 1.050 ± 0.068 mg mL−1 (k = 2), respectively. [ABSTRACT FROM AUTHOR]

Copyright of Journal of Food Safety & Quality is the property of Journal of Food Safety & Quality Editorial Department and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Fermentation is a preservation process responsible for increasing food product shelf life. In this context, alcoholic fermentation can add value to unconventional Amazon fruits, e.g., araçá-boi (Eugenia stipitata). This fruit has various antioxidant phenolic compounds with well-known nutraceutical properties. However, araçá-boi is still underexplored by food industry. This rationale led to investigate the influence of five commercial yeasts (Saccharomyces cerevisiae) and filtration process on chemical composition and antioxidant capacity of araçá-boi beverages. DPPH and Folin Ciocalteu assays were used to determine antioxidant capacity and total phenolic content. Organic compounds' contents were assessed by NMR-ERETIC2. In all beverages, ten compounds [tyrosol, sucrose, fructose, (α/β)-glucose, ethanol, malic, citric, gallic, and succinic acids] were identified and quantified. The highest phenolic concentrations [gallic acid (390.0 µM) and tyrosol (380.0 µM)] were found in Biolievito Bayanus (BBA) beverage. The new BBA beverage was used for investigating filtration process influence on chemical composition and antioxidant responses. Alcoholic content (unfiltered: 13.9°GL and filtered: 12.7°GL), antioxidant responses, and total phenolic contents were influenced by filtration process. The yeast type and unfiltered process were determinant for chemical content and antioxidant capacity of beverages. These results might be useful to private sector and future production and commercialization of araçá-boi beverages. [ABSTRACT FROM AUTHOR]

A systematic procedure for the determination of purity values of amino acid reference materials was developed by use of mass balance method where four categories of impurities (related structure impurities (RSIs), water, organic solvent residue (OSR), and non-volatile residue (NVR)) were quantified separately. The amount of RSIs was determined using a combination of three quantification methods. To ensure metrological traceability in the determination of RSIs, at least one such impurity in each candidate amino acid reference material was quantified using liquid chromatography–isotope dilution tandem mass spectrometry (LC-IDMS/MS). Other RSIs were determined using external calibration liquid chromatography–tandem mass spectrometry (LC-MS/MS) or o-phthaldialdehyde (OPA) derivatization, followed by liquid chromatography–ultraviolet (LC-UV) measurement. As the UV absorption of most RSIs came basically from the same chromophore after OPA derivatization, a relative peak area approach was used in the LC-UV method to quantify the amount of RSIs by comparing their peak areas with that of a reference RSI. The reference RSI was pre-selected and the amount determined by LC-IDMS/MS separately. The absence of D-amino acids was confirmed using Marfey's reagent derivatization, followed by LC-UV analysis. The amounts of water, OSR, and NVR were measured using Karl Fischer coulometry, gas chromatography–mass spectrometry (GC-MS) and thermogravimetry, respectively. By using this procedure, four amino acid (L-valine, L-leucine, L-isoleucine, and L-phenylalanine) certified reference materials (CRMs) were developed from the candidate materials. The homogeneity and stability of the CRMs were demonstrated by use of LC-IDMS/MS or OPA-LC-UV method, following the principles in ISO 17034 and ISO Guide 35. Graphical abstract [ABSTRACT FROM AUTHOR]

Whey protein-based supplements are one of the most consumed products to improve performance, and they are, generally, recognized as being safe. However, recent studies indicated different types of adulterations in these products, including the presence of diuretics and stimulants, allowing the possibility of contamination by other substances with pharmacological activities, such as anabolic agents. In this sense, the aim of this study was to develop and validate a method for the screening of 46 anabolic agents in whey protein. The analytical method consisted of sample preparation using liquid-liquid extraction, followed by analytes derivatization and, finally, analysis by gas chromatography coupled to triple quadrupole mass spectrometry in two different experiments: multiple reaction monitoring and full scan. After validation, the method proved to be specific, accurate, and precise; no carry over effect was observed, and the limit of detection values varied between 0.5 and 100 ng g−1. The analysis of 44 commercial samples did not detect the presence of any anabolic agent. However, full scan analysis detected caffeine in 14 samples and bisphenol A in one of them. Therefore, these results indicate that the analyzed samples are safe concerning the presence of the evaluated anabolic agents, but it does not mean that they are free from the presence of other substances with pharmacological activities which are not declared on label. [ABSTRACT FROM AUTHOR]

Background: Human urine gives evidence of the metabolism in the body and contains different metabolites at various concentrations. A number of analytical techniques including mass spectrometry (MS) and nuclear magnetic resonance (NMR) have been used to obtain metabolites levels in urine samples. However, gas chromatography–mass spectrometry (GC–MS) is one of the most widely used techniques for urinary metabolomics studies due to its higher sensitivity, resolution, reproducibility, reliability, relatively low cost and ease of operation compared to liquid chromatography–mass spectrometry and NMR. Aim of Review: This review looks at various aspects of urine preparation prior to analysis by GC–MS including sample storage, urease pretreatment, derivatization, use of internal standard and quality control samples for data correction. In addition, most common types of inlet liners, ionization techniques and columns are discussed and a summary of mass analyzers are also highlighted. Lastly, the role of retention index in metabolite identification and data normalization methods are presented. Key scientific concepts of review: The purpose of this review is summarizing methods of sample storage, pretreatment, and GC–MS analysis that are mostly used in urine metabolomics studies. Specific emphasis is given to the critical steps within the GC–MS urine metabolomics that those new to this field need to be aware of and the remaining challenges that require further attention and studies. [ABSTRACT FROM AUTHOR]

Salvianolic acid A (Sal A), a water-soluble ingredient in Danshen, has various biological activities. Sal A and its impurities have similar physical and chemical properties, as well as strong reducibility; therefore, they are difficult to prepare and purify. In this study, high-purity Sal A was obtained by purification of sephadex chromatography and preparative chromatography. Furthermore, HPLC–DAD tandem ECD and HPLC–DAD tandem MS methods were used for non-volatile organic impurity analysis, ICP-MS method was used for non-volatile inorganic impurities and mass balance method and quantitative nuclear magnetic resonance were employed to certify the product. The structures of Sal A and its relative impurities were validated by nuclear magnetic resonance spectroscopy and mass spectrometry, and their contents were quantified as well. Following the principles of ISO Guides 34:2009 and 35:2005, a Sal A reference material was certified, covering homogeneity studies, stability studies, characterization, and uncertainty estimations. [ABSTRACT FROM AUTHOR]

Metrological traceability for organic chemical measurements is a documented unbroken chain of calibrations with stated uncertainties that ideally link the measurement result for a sample to a primary calibrator in appropriate SI units (e.g., mass fraction). A comprehensive chemical purity determination of the organic calibrator is required to ensure a true assessment of this result. We explore the evolution of chemical purity capabilities across metrology institute members of the Consultative Committee for Amount of Substance: Metrology in Chemistry and Biology's Organic Analysis Working Group (OAWG). The OAWG work program has promoted the development of robust measurement capabilities, using indirect "mass balance" determinations via rigorous assessment of impurities and direct determination using quantitative nuclear magnetic resonance spectroscopy methods. A combination of mass balance and qNMR has been shown to provide a best practice approach. Awareness of the importance of the traceability of organic calibrators continues to grow across stakeholder groups, particularly in key areas such as clinical chemistry where activities related to the Joint Committee for Traceability in Laboratory Medicine have raised the profile of traceable calibrators. [ABSTRACT FROM AUTHOR]

The anabolic‐androgenic steroid methylstenbolone (MSTEN; 2α,17α‐dimethyl‐17β‐hydroxy‐5α‐androst‐1‐en‐3‐one) is available as a so‐called designer steroid or nutritional supplement. It is occasionally detected in doping control samples, predominantly tested and confirmed as the glucuronic acid conjugate of methylstenbolone. The absence of other meaningful metabolites reported as target analytes for sports drug testing purposes can be explained by the advertised metabolic stability of methylstenbolone. In 2013, a first investigation into the human metabolism of methylstenbolone was published, and two hydroxylated metabolites were identified as potential targets for initial testing procedures in doping controls. These metabolites were not observed in recent doping control samples that yielded adverse analytical findings for methylstenbolone, and in the light of additional data originating from a recent publication on the in vivo metabolism of methylstenbolone in the horse, revisiting the metabolic reactions in humans appeared warranted. Therefore, deuterated methylstenbolone together with hydrogen isotope ratio mass spectrometry (IRMS) in combination with high accuracy/high resolution mass spectrometry were employed. After oral administration of a single dose of 10 mg of doubly labeled methylstenbolone, urine samples were collected for 29 days. Up to 40 different deuterated methylstenbolone metabolites were detected in post‐administration samples, predominantly as glucuronic acid conjugates, and all were investigated regarding their potential to prolong the detection window for doping controls. Besides methylstenbolone excreted glucuronidated, three additional metabolites were still detectable at the end of the study on day 29. The most promising candidates for inclusion into routine sports drug testing methods (2α,17α‐dimethyl‐5α‐androst‐1‐ene‐3β,17β‐diol and 2α,17α‐dimethyl‐5α‐androst‐1‐ene‐3α,17β‐diol) were synthesized and characterized by NMR. [ABSTRACT FROM AUTHOR]

Copyright of Colombia Internacional is the property of Universidad de los Andes and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Copyright of International Journal of Endocrinology / Mìžnarodnij Endokrinologìčnij Žurnal is the property of Zaslavsky O.Yu and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Copyright of Journal of Food Safety & Quality is the property of Journal of Food Safety & Quality Editorial Department and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

By using post-column reaction GC, a method for accurate and direct characterization of musty odor substances (2-methylisoborneol and geosmin) in a candidate standard solution has been developed. Because the GC system gives a response depending on the number of carbon atoms constituting the organic compound by combustion and conversion to methane, naphthalene and diethyl phthalate, purity of which had been accurately assessed, were used as reference materials for calibration. To verify the method, concentrations of 2-methylisoborneol and geosmin in the sample determined by the method were compared with the concentrations calculated based on gravimetric mixing that is one of the general methods for the characterization of the organic standard solutions. The analytical results of each analyte obtained by the two methods agreed within a deviation of about 0.5 % or less. [ABSTRACT FROM AUTHOR]

This is a first look at a non‐targeted screening method based on Orbitrap gas chromatography–mass spectrometry (GC–MS) technology for a large number of banned compounds in sports found in urine, including exogenous anabolic steroids, β‐agonists, narcotics, stimulants, hormone modulators, and diuretics. A simple sample preparation was processed in four steps: an enzymatic hydrolysis, liquid–liquid extraction, evaporation, and trimethylsilylation. All compounds were able to meet the World Anti‐Doping Agency's sensitivity criteria with mass accuracies less than 1 ppm and with sufficient points across the peak by running the Orbitrap GC–MS in full‐scan mode. In addition, we discuss our initial findings of using a full‐scan selected ion monitoring‐tandem mass spectrometry (SIM‐MS/MS) approach as a way to obtain lower detection limits and reach desirable selectivity for some exogenous anabolic steroids. [ABSTRACT FROM AUTHOR]

Due to the internationalisation of food production and distribution, there has been a significant increase of food fraud in recent years. Food fraud can have serious health implications, and it occurs when food manufacturers implement unethical practices such as making false label claims as well as using additives and fillers within their products to increase profitability. This has been a serious concern. Meat adulteration was examined using a well-defined proteogenomic annotation, carefully selected surrogate tryptic peptides and high-resolution mass spectrometry. Selected mammalian meat samples were homogenised and the proteins extracted and digested with trypsin. Chromatography was achieved using a 30-min linear gradient along with a BioBasic C8 100 × 1 mm column at a flow rate of 75 µl min–1. The mass spectrometer was operated in full-scan high-resolution and accurate mass using resolving powers of 140,000 and 17,500 (FWHM) in full-scan MS and MS/MS respectively. Data independent acquisition (DIA) mode was used including 12 DIA MS/MS scans to cover the mass range 600–1200m/z. Methodicallyin silicoanalyses of myoglobin, myosin-1, myosin-2 and β-haemoglobin sequences allow for the identification of a species-specific tryptic peptide mass lists and theoretical MS/MS spectra. Following comprehensive MS, MS/MS or DIA analyses, the method was capable of the detection and identification of very specific tryptic peptides for all four targeted proteins for each animal species tested with observedm/zbelow 3 ppm compared with the theoreticalm/z. The analyses were successfully performed with raw and cooked meat. Specifically, the method was capable of detecting 1% (w/w) of pork or horse meat in a mixture before and after cooking (71°C internal temperature). [ABSTRACT FROM AUTHOR]

The power of foodomics as a discipline that is now broadly used for quality assurance of food products and adulteration identification, as well as for determining the safety of food, is presented. Concerning sample preparation and application, maintenance of highly sophisticated instruments for both high-performance and high-throughput techniques, and analysis and data interpretation, special att ention has to be paid to the development of skilled analysts. The obtained data shall be integrated under a strong bioinformatics environment. Modern mass spectrometry is an extremely powerful analytical tool since it can provide direct qualitative and quantitative information about a molecule of interest from only a minute amount of sample. Quality of this information is influenced by the sample preparation procedure, the type of mass spectrometer used and the analyst's skills. Technical advances are bringing new instruments of increased sensitivity, resolution and speed to the market. Other methods presented here give additional information and can be used as complementary tools to mass spectrometry or for validation of obtained results. Genomics and transcriptomics, as well as affinity-based methods, still have a broad use in food analysis. Serious drawbacks of some of them, especially the affinity-based methods, are the cross-reactivity between similar molecules and the influence of complex food matrices. However, these techniques can be used for pre-screening in order to reduce the large number of samples. Great progress has been made in the application of bioinformatics in foodomics. These developments enabled processing of large amounts of generated data for both identification and quantification, and for corresponding modeling. [ABSTRACT FROM AUTHOR]

The use of anabolic androgenic steroids (AAS) has grown among practitioners of recreational bodybuilding, with significant contributions of designer steroids, aiming muscle hypertrophy in healthy subjects. The abusive use of AAS in general is associated with adverse effects; one of the most worrisome is cancer development. The aim of this study was to evaluate the effectiveness of the cytokinesis block micronucleus (CBMN) test in human lymphocytes in identifying risk groups for cancer development in users of AAS. Blood was collected from 15 AAS users bodybuilders (G1), 20 non-users bodybuilders (G2) and 20 non-users sedentary (G3). MN analysis was performed on a minimum of 1000 binucleated lymphocytes. The occurrence of MN was significantly higher (p < 0.05) in individuals of G1 compared to G2 and G3. The results indicate the sensitivity of CBMN in human lymphocytes in the identification of chromosomal damage in consequence of AAS. [ABSTRACT FROM AUTHOR]

N, N-dimethyl-2-phenylpropan-1-amine (NN-DMPPA) is a new designer stimulant prohibited in sport in-competition according to the List of Prohibited Substances and Methods published by the World Anti-Doping Agency (WADA). The first published data on the excretion study of NN-DMPPA to support the knowledge of NN-DMPPA in routine anti-doping control have been presented. The reliable gas chromatography-mass spectrometry quantitative method (GC-MS) has been validated and applied to the excretion study of NN-DMPPA. The validation parameters of the GC-MS method for determination of NN-DMPPA in human urine were the linear calibration range of 100 to 7500 ng/mL, the LOD of 13.9 ng/mL and the LOQ of 42.2 ng/mL. According to the obtained repeatability, intermediate precision, and trueness, the applied GC-MS method was precise and accurate. Urine samples from three volunteers in the excretion study were collected for 5 days after single oral administration of the supplement NOXPUMP containing NN-DMPPA. The obtained results showed the maximum concentration of NN-DMPPA (189-303 ng/mL) in urine samples at a time of 2-3 h post-administration. The NN-DMPPA concentration in urine samples was higher than 50 ng/mL until 22-23 h after the dietary supplement ingestion. This means that according to the WADA rules the use of a supplement containing NN-DMPPA may be related to a positive case when athletes took this supplement in-competition. Moreover, excretion results demonstrate also that NN-DMPPA may be detected in urine samples by the applied GC-MS method till 46 h after supplement administration. Additionally, the excretion study of β-methylphenethylamine as the second prohibited substance present in the supplement NOXPUMP has been investigated. [Figure not available: see fulltext.] [ABSTRACT FROM AUTHOR]

Salbutamol (SAL) is the most widely used β2-agonist drug for asthma and chronic obstructive pulmonary patients, but it is also often abused as feed additive. In recent years, the abuse of SAL has led to a large number of food safety incidents. Therefore, the monitoring of SAL residues in animal products is very important. A highly selective boronate affinity magnetic adsorbent was synthesized and developed for detection of trace levels of SAL residues in pig tissue samples. The obtained Fe3O4@SiO2@FPBA(4-formylphenylboronic acid) magnetic adsorbent showed good adsorption ability to catechol and SAL, and then it was successfully applied as special magnetic solid-phase phase extraction adsorbent coupled with high-performance liquid chromatography (HPLC) for simultaneous isolation and determination of cis-diol compounds. The binding capacity of catechol and SAL reached 96 and 50 µmol/g, respectively. The method was successfully established for the detection of trace levels of SAL in pig tissue samples. The linear range extended from 0.32 to 800 µg/kg ( R2 = 0.9994). The limit of detection of SAL was 0.19 µg/kg. The recoveries were satisfactory (89.5-108.0%) at three spiked levels with RSD between 2.1 and 11.3%. These results indicated that the method has potential for enrichment and detection of trace levels of SAL residual in animal food products. Copyright © 2015 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Quantitative NMR spectroscopy (qNMR) has been examined for purity assessment using a range of organic calibration standards of varying structural complexities, certified using the traditional mass balance approach. Demonstrated equivalence between the two independent purity values confirmed the accuracy of qNMR and highlighted the benefit of using both methods in tandem to minimise the potential for hidden bias, thereby conferring greater confidence in the overall purity assessment. A comprehensive approach to purity assessment is detailed, utilising, where appropriate, multiple peaks in the qNMR spectrum, chosen on the basis of scientific reason and statistical analysis. Two examples are presented in which differences between the purity assignment by qNMR and mass balance are addressed in different ways depending on the requirement of the end user, affording fit-for-purpose calibration standards in a cost-effective manner. [ABSTRACT FROM AUTHOR]

A simple, sensitive and specific liquid chromatography-tandem mass spectrometry method was developed and validated for quantification of exemestane. Exemestane is an irreversible, steroidal aromatase inactivator and it is indicated for adjuvant treatment of postmenopausal women with estrogen-receptor positive early breast cancer. The analyte and internal standard (IS)(exemestane-D2) were extracted by liquid-liquid extraction withethyl acetate: n-Hexane (80:20v/v) solvent mixture. The chromatographic separation was performed on a reverse phase Kinetex 2.6μ C18, 50 × 4.6 mm column with a mobile phase of 0.01% (v/v) acetic acid in water andacetonitrilein gradient composition. Exemestane was quantitated in positive ionization by multiple-reaction-monitoring with a mass spectrometer. The mass transitions m/z 297.4 → 121.2 and m/z 299.4 → 123.2 were used to measure exemestane and exemestane-D2 respectively. The assay exhibited a linear dynamic range of 0.100-40.0 ng/ml for exemestane in human plasma. The lower limit of quantitation achieved was 0.100ng/mL for exemestanewith a relative standard deviation of less than 20%. Acceptable precision and accuracies were obtained for concentrations over the standard curve ranges.A run time of 8.0 minutes for each sample made it possible to analyze more than 100 human plasma samples per day. The validated method has been successfully used to analyze human plasma samples for application in pharmacokinetic and bioequivalence studies. [ABSTRACT FROM AUTHOR]

Colloquially referred to by various misleading monikers ('pro-hormones', 'natural steroids', 'testosterone boosters', etc.) designer anabolic steroids have been popular now for over a decade as a way to achieve classic anabolic steroid-like results from products sold in the legal marketplace. Recent evidence suggests that anabolic steroid use may be the most common cause of hypogonadism in men of reproductive age. Despite recent regulatory efforts that have banned specific compounds, many anabolic-androgenic steroids ( AAS) remain available in over-the-counter dietary supplements that are legally sold in the United States. Severe side effects including hepatotoxicity, cholestasis, renal failure, hypogonadism, gynecomastia, and infertility have been reported secondary to the use of these products. While some of these side effects may be reversible, more aggressive use may result in more permanent end-organ damage as has been previously described for the case of aggressive AAS users (Rahnema et al., Fertil Steril, 2014). Designer AAS remain easily available for purchase in over-the-counter bodybuilding supplements and these products appear to be increasingly popular, despite the known health risks associated with their use. We conducted a systematic search to identify the designer steroids that are most commonly sold in dietary supplements as of April 2014 and review what is known regarding their potency and toxicity. We propose that the impact of AAS use on the reproductive and hormonal health of men is underestimated in the literature owing to previous studies' failure to account for designer steroid use. Lastly, we make clinical recommendations to help physicians steer patients away from potentially harmful supplements, and summarize key regulatory obstacles that have allowed potent androgens to remain unregulated in the legal marketplace. [ABSTRACT FROM AUTHOR]

ABSTRACT Anti-doping laboratories need to be aware of evolutions on the steroid market and elucidate steroid metabolism to identify markers of misuse. Owing to ethical considerations, in vivo and in vitro models are preferred to human excretion for nonpharmaceutical grade substances. In this study the chimeric mouse model and human liver microsomes (HLM) were used to elucidate the phase I metabolism of a new steroid product containing, according to the label, methylstenbolone. Analysis revealed the presence of both methylstenbolone and methasterone, a structurally closely related steroid. Via HPLC fraction collection, methylstenbolone was isolated and studied with both models. Using HLM, 10 mono-hydroxylated derivatives (U1-U10) and a still unidentified derivative of methylstenbolone (U13) were detected. In chimeric mouse urine only di-hydroxylated metabolites (U11-U12) were identified. Although closely related, neither methasterone nor its metabolites were detected after administration of isolated methylstenbolone. Administration of the steroid product resulted mainly in the detection of methasterone metabolites, which were similar to those already described in the literature. Methylstenbolone metabolites previously described were not detected. A GC-MS/MS multiple reaction monitoring method was developed to detect methylstenbolone misuse. In one out of three samples, previously tested positive for methasterone, methylstenbolone and U13 were additionally detected, indicating the applicability of the method. Copyright © 2014 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Herbal and nutritional supplements are more and more popular in the western population. One of them is an extract of an exotic plant, named Tribulus terrestris (TT). TT is a component of several supplements that are available over-the-counter and widely recommended, generally as enhancers of human vitality. TT is touted as a testosterone booster and remedy for impaired erectile function; therefore, it is targeted at physically active men, including male athletes. Based on the scientific literature describing the results of clinical trials, this review attempted to verify information on marketing TT with particular reference to the needs of athletes. It was found that there are few reliable data on the usefulness of TT in competitive sport. In humans, a TT extract used alone without additional components does not improve androgenic status or physical performance among athletes. The results of a few studies have showed that the combination of TT with other pharmacological components increases testosterone levels, but it was not discovered which components of the mixture contributed to that effect. TT contains several organic compounds including alkaloids and steroidal glycosides, of which pharmacological action in humans is not completely explained. One anti-doping study reported an incident with a TT supplement contaminated by a banned steroid. Toxicological studies regarding TT have been carried out on animals only, however, one accidental poisoning of a man was described. The Australian Institute of Sport does not recommend athletes' usage of TT. So far, the published data concerning TT do not provide strong evidence for either usefulness or safe usage in sport. [ABSTRACT FROM AUTHOR]

Plant-based protein powders are rapidly growing in popularity, and outdated quality assurance tools expose vulnerabilities to adulteration via different methods of "protein spiking". Adequate diagnostic tools are urgently needed to be able to authenticate protein source ingredients and screen for potential adulterants. We explored the application of three diagnostic tools for ingredient identification: targeted PCR with Sanger sequencing, NGS, and LC-MS/MS. We collected 33 samples of common commercial products from the plant-based protein powder market and sought to identify botanical components using the three technologies. We found success in detection with all approaches, with at least one main protein source being identified by at least one approach in all samples. The investigation uncovered challenges to data collection or result interpretation with each technology including but not limited to amplification biases with PCR technologies, potential influence of DNA degradation, and issues with protein solubility during isolation. Ultimately, each platform demonstrated utility along with certain caveats, which epitomized the importance of orthogonality of testing. [ABSTRACT FROM AUTHOR]