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Analytical and Bioanalytical Chemistry Jul 2015A method for the determination and quantification of ketosteroid hormones in meat by mass spectrometry, based on the derivatization of the carbonyl moiety of steroids by...
A method for the determination and quantification of ketosteroid hormones in meat by mass spectrometry, based on the derivatization of the carbonyl moiety of steroids by O-methylhydroxylamine, is presented. The quantitative assay is performed by means of multiple-reaction-monitoring (MRM) scan mode and using the corresponding labelled species, obtained by reaction with d 3-methoxylamine, as internal standard. The accuracy of the method was established by evaluating artificially spiked samples, obtaining values in the range 90-110%. Recovery tests were performed on blank matrix samples spiked with non-natural steroids including trenbolone and melengestrol acetate. The latter experiment revealed that the yield of the extraction processes was approximately 60%. Good values of LOQ and LOD were achieved, making this method competitive with current hormone assay methods.
Topics: Ketosteroids; Meat; Solid Phase Microextraction; Tandem Mass Spectrometry
PubMed: 26014285
DOI: 10.1007/s00216-015-8772-5 -
Journal of Chromatography. B,... Apr 2015An ultra-performance convergence chromatography (UPC2) system coupled tandem mass spectrometry was successfully utilised to analyse chlormadinone acetate, delmadinone...
An ultra-performance convergence chromatography (UPC2) system coupled tandem mass spectrometry was successfully utilised to analyse chlormadinone acetate, delmadinone acetate, fluorogestone acetate, medroxyprogesterone acetate, megestrol acetate, melengestrol acetate, chlortestasterone acetate in bovine and porcine kidney fat. This novel approach obtained an improved resolution in comparison to previously reported chromatographic methods combined with MS detector in a shorter analytical time. All the acetylgestagen compounds were well separated on a ACQUITY UPC(2) HSS C18 column (3.0 × 100 mm, 1.7 μm) by applying methanol and carbon dioxide (2/98). The LOQ of delmadinone acetate, melengestrol acetate, medroxyprogesterone acetate and megestrol acetate are 0.5 μg/kg, fluorogestone acetate, chlormadinone acetate and chlortestasterone acetate 1.0 μg/kg. The recoveries of gestagens spiked in kidney fats at a concentration range of 0.5 to 4 μg/kg were above 86.1% with relative standard deviations (RSD) less than 13.1%. These rapid and reliable methods can be used to efficiently separate, characterize and quantify the residues of gestagens in kidney fats with advantages of shorter time, more sensitive and environmental friendly.
Topics: Adipose Tissue; Animals; Cattle; Chromatography, High Pressure Liquid; Kidney; Linear Models; Progestins; Reproducibility of Results; Sensitivity and Specificity; Swine; Tandem Mass Spectrometry
PubMed: 25777477
DOI: 10.1016/j.jchromb.2015.02.034 -
Journal of Agricultural and Food... Dec 2014This work examines the fate of synthetic growth promoters (trenbolone acetate, melengestrol acetate, and zeranol) in sterilized soil systems, focusing on their sorption...
This work examines the fate of synthetic growth promoters (trenbolone acetate, melengestrol acetate, and zeranol) in sterilized soil systems, focusing on their sorption to organic matter and propensity for mineral-promoted reactions. In organic-rich soil matrices (e.g., Pahokee Peat), the extent and reversibility of sorption did not generally correlate with compound hydrophobicity (e.g., K(ow) values), suggesting that specific binding interactions (e.g., potentially hydrogen bonding through C17 hydroxyl groups for the trenbolone and melengestrol families) can also contribute to uptake. In soils with lower organic carbon contents (1-5.9% OC), evidence supports sorption occurring in parallel with surface reaction on inorganic mineral phases. Subsequent experiments with pure mineral phases representative of those naturally abundant in soil (e.g., iron, silica, and manganese oxides) suggest that growth promoters are prone to mineral-promoted oxidation, hydrolysis, and/or nucleophilic (e.g., H2O or OH(-)) addition reactions. Although reaction products remain unidentified, this study shows that synthetic growth promoters can undergo abiotic transformation in soil systems, a previously unidentified fate pathway with implications for their persistence and ecosystem effects in the subsurface.
Topics: Adsorption; Growth Hormone; Kinetics; Melengestrol Acetate; Minerals; Oxidation-Reduction; Soil; Soil Pollutants; Trenbolone Acetate; Zeranol
PubMed: 25426694
DOI: 10.1021/jf5035527 -
Se Pu = Chinese Journal of... Jun 2014A new method using TurboFlow on-line cleanup liquid chromatography combined with tandem mass spectrometry (MS/MS) has been developed for simultaneous determination of...
A new method using TurboFlow on-line cleanup liquid chromatography combined with tandem mass spectrometry (MS/MS) has been developed for simultaneous determination of six progesterones including 19-norethindrone, 17alpha-hydroxyprogesterone, medroxyprogesterone, megestrol acetate, progesterone and melengestrol acetate in milk. Samples were extracted by acetonitrile. The analytes in extract were on-line purified directly on Cyclone-P purification column where the sample matrices were washed away. Subsequently, the analytes which were eluted from the extraction column onto Phenyl-Hexyl column were separated with a gradient elution, and detected with electrospray ionization mass spectrometry in the positive scan mode using multiple reaction monitoring (MRM). The isotope internal standards were employed for quantification. As a result, the linearities were satisfactory with the correlation coefficients of > 0.999 at concentrations ranging from 0.1 microg/L to 50 microg/L. Based on the repeated analysis of a known blank sample, the limit of quantification (LOQ) is 0.5 microg/kg. Average recoveries of the analytes fortified at three levels (1, 5 and 25 microg/kg) ranged from 90.8% to 107.5%, and the relative standard deviations (RSDs) ranged from 6.3% to 11.8%. This proposed method is simple, rapid, sensitive and highly selective, and can be applied to simultaneous identification an quantification of the six progesterones in milk.
Topics: Animals; Chromatography, Liquid; Milk; Progesterone; Spectrometry, Mass, Electrospray Ionization; Tandem Mass Spectrometry
PubMed: 25269265
DOI: 10.3724/sp.j.1123.2014.01044 -
Steroids Aug 2014Biotransformation of melengestrol acetate (MGA, 17α-acetoxy-6-methyl-16-methylenepregna-4,6-diene-3,20-dione) (1) was investigated for the first time by using fungal...
Biotransformation of melengestrol acetate (MGA, 17α-acetoxy-6-methyl-16-methylenepregna-4,6-diene-3,20-dione) (1) was investigated for the first time by using fungal cultures. Incubation of compound 1 with Cunninghamella blakesleeana yielded a new major metabolite, 17α-acetoxy-11β-hydroxy-6-methyl-16-methylenepregna-4,6-diene-3,20-dione (2). The metabolite 2 was purified by using HPLC, followed by characterization through (1)H- and (13)C-NMR and other spectroscopic techniques. Single crystal X-ray diffraction analysis was used to deduce the three dimensional structures of melengestrol acetate (1) and metabolite 2 for the first time. T-cell proliferation assay was employed to evaluate the immunosuppressant effect of compounds 1 and 2 with IC50=0.5±0.07 and 0.6±0.08μg/mL, respectively. The results indicated that these compounds possess sixfold potent T-cell proliferation inhibitory activity as compared to the standard prednisolone (IC50<3.1μg/mL). Both compounds were found to be non-toxic in a 3T3 (mouse fibroblast) cell-based cytotoxicity assay. This discovery of potent anti-inflammatory activity of compounds 1 and 2 can lead the way to develop new immunosuppressant compounds for clinical application.
Topics: 3T3 Cells; Animals; Anti-Inflammatory Agents, Non-Steroidal; Biotransformation; Cell Proliferation; Cunninghamella; Dose-Response Relationship, Drug; Immunosuppressive Agents; Melengestrol Acetate; Mice; Models, Molecular; Molecular Conformation; Structure-Activity Relationship; T-Lymphocytes
PubMed: 24793568
DOI: 10.1016/j.steroids.2014.04.012 -
Food Additives & Contaminants. Part A,... Apr 2014Levels of several natural urinary steroids have been determined in the urine of a large number of animals of different cattle categories in the context of steroid abuse...
Levels of several natural urinary steroids have been determined in the urine of a large number of animals of different cattle categories in the context of steroid abuse in beef production. Bovine animals of different breeds, sex and age included in the Slovene national residue detection plan for steroid abuse were studied. Urine from 120 males and 174 females was analysed. Urinary boldenone, boldione, androstenedione, equiline, medroxyprogesterone, medroxyprogesterone acetate, melengestrol acetate, progesterone, stanozolol, trenbolone, trenbolone acetate, 17α-ethinylestradiol, 17α-methyltestosterone, epitestosterone, 17β-estradiol, testosterone, and nandrolone were determined by LC-MS/MS. Epitestosterone was found in all bulls; while the proportion of animals containing testosterone and androstenedione increased with age. Testosterone was not detected in bulls less than 5 months of age. Epitestosterone levels, however, were not age dependent. The ratio of testosterone to epitestosterone thus increased with age, from 0.13 ± 0.09 at 1-7 months to 0.42 ± 0.10 at 25-38 months. It was significantly (p < 0.01) higher in bulls above 13 months than in younger animals. In contrast to males, no urinary testosterone was found in females, whereas epitestosterone, androstenedione, progesterone and estradiol were present. The proportion of animals of various age groups in which epitestosterone was detected ranged from 68% to 100%, but the differences were not significant. The presence of both estradiol and progesterone in the same sample was not observed in any animal. The results of this study could be helpful in determining physiological urinary steroid levels in order to provide a baseline for the control of steroid abuse in beef production.
Topics: Aging; Animals; Cattle; Female; Gonadal Steroid Hormones; Male
PubMed: 24405322
DOI: 10.1080/19440049.2013.880000 -
Antimicrobial Agents and Chemotherapy 2014Candida species are the cause of 60% of all mycoses in immunosuppressed individuals, leading to ∼150,000 deaths annually due to systemic infections, whereas the...
Candida species are the cause of 60% of all mycoses in immunosuppressed individuals, leading to ∼150,000 deaths annually due to systemic infections, whereas the current antifungal therapies either have toxic side effects or are insufficiently efficient. We performed a screening of two compound libraries, the Enzo and the Institute for Molecular Medicine Finland (FIMM) oncology collection library, for anti-Candida activity based on the European Committee on Antimicrobial Susceptibility Testing (EUCAST) guidelines. From a total of 844 drugs, 26 agents showed activity against Candida albicans. Of those, 12 were standard antifungal drugs (SADs) and 7 were off-target drugs previously reported to be active against Candida spp. The remaining 7 off-target drugs, amonafide, tosedostat, megestrol acetate, melengestrol acetate, stanozolol, trifluperidol, and haloperidol, were identified with this screen. The anti-Candida activities of the new agents were investigated by three individual assays using optical density, ATP levels, and microscopy. The antifungal activities of these drugs were comparable to those of the SADs found in the screen. The aminopeptidase inhibitor tosedostat, which is currently in a clinical trial phase for anticancer therapy, displayed a broad antifungal activity against different Candida spp., including Candida glabrata. Thus, this screen reveals agents that were previously unknown to be anti-Candida agents, which allows for the design of novel therapies against invasive candidiasis.
Topics: Antifungal Agents; Antineoplastic Agents; Candida; Clinical Trials as Topic; Drug Discovery; Drug Repositioning; Drug Resistance, Fungal; Glycine; High-Throughput Screening Assays; Humans; Hydroxamic Acids; Microbial Sensitivity Tests; Small Molecule Libraries
PubMed: 24277040
DOI: 10.1128/AAC.01087-13 -
Food Additives & Contaminants. Part A,... 2013A method for the determination of residues of six steroids with gestagenic action (altrenogest, medroxyprogesterone acetate, megestrol acetate, melengestrol acetate,...
A method for the determination of residues of six steroids with gestagenic action (altrenogest, medroxyprogesterone acetate, megestrol acetate, melengestrol acetate, acetoxyprogesterone and chlormadinone acetate) in animal fat tissue was developed. The procedure consists of methanol extraction, clean-up on an alumina column and LC-MS/MS measurement. The method has been validated according to Decision 2002/657/EC. Decision limits for all analytes were observed within the range from 0.3 to 1.7 ng g(-1), and recoveries were between 80% and 105%. The method is robust and can be applied for both screening and confirmatory analyses in routine residue monitoring.
Topics: Adipose Tissue; Animals; Chromatography, Liquid; Kidney; Progestins; Reproducibility of Results; Tandem Mass Spectrometry
PubMed: 23710547
DOI: 10.1080/19440049.2013.789555 -
Regulatory Toxicology and Pharmacology... Apr 2013Some synthetic chemicals are suspected to be responsible for adverse effects on endocrine function. Sex hormones administered to farm animals are of particular interest...
Plasma concentrations of melengestrol acetate in humans extrapolated from the pharmacokinetics established in in vivo experiments with rats and chimeric mice with humanized liver and physiologically based pharmacokinetic modeling.
Some synthetic chemicals are suspected to be responsible for adverse effects on endocrine function. Sex hormones administered to farm animals are of particular interest because of their regulatory role in developmental processes. To predict concentrations in humans of the synthetic growth promoter melengestrol acetate (17α-acetoxy-6-methyl-16-methylenepregna-4,6-diene-3,20-dione), a forward dosimetry approach was carried out using data from no-observed-adverse-effect-level doses orally administered to mice or rats and from in vitro human and rodent experiments. Human liver microsomes preferentially mediated 2-hydroxylation of melengestrol acetate, but rodent livers produced additional unidentified hydroxymetabolites. Adjusted animal biomonitoring equivalents for melengestrol acetate from mouse and rat studies were scaled to human biomonitoring equivalents using known species allometric scaling factors and human metabolic data with a simple physiologically based pharmacokinetic (PBPK) model. Melengestrol acetate elimination in humans was estimated to be slow compared with elimination in rodents. The disposition of melengestrol acetate in humans was evaluated using chimeric TK-NOG mice with humanized liver. The results suggest the usefulness of simplified PBPK modeling combined with in vitro and in vivo experiments and literature resources as well as a future interest in estimating by a full PBPK modeling using another bottom up system. This model may also be useful for risk evaluation and for simulating plasma concentrations resulting from exposure to low doses of melengestrol acetate and related compounds.
Topics: Animals; Cells, Cultured; Chimera; Glucocorticoids; Hepatocytes; Humans; Liver; Male; Melengestrol Acetate; Mice; Mice, Transgenic; Microsomes, Liver; Models, Biological; No-Observed-Adverse-Effect Level; Rats; Rats, Sprague-Dawley
PubMed: 23395687
DOI: 10.1016/j.yrtph.2013.01.008 -
Reproduction in Domestic Animals =... Dec 2012North American zoos began using melengestrol acetate (MGA) implants to control reproduction in wild felids in the mid-1970s. Research linking MGA and other...
North American zoos began using melengestrol acetate (MGA) implants to control reproduction in wild felids in the mid-1970s. Research linking MGA and other progestin-based contraceptives to uterine and mammary pathology in canids as well as felids resulted in a shift to GnRH agonist implants (Suprelorin(®): deslorelin, Peptech Animal Health, Australia). However, a recent study revealed an association between Suprelorin(®) and uterine pathology in canids, but that pathology was not found in canids treated with oral megestrol acetate (MA) for 2 weeks around the time of implant insertion to prevent the initial agonist stimulation phase. Thus, the AZA Wildlife Contraception Center (WCC) currently recommends Suprelorin(®) plus the 2-week MA regimen for wild canids and felids. WCC research is now focusing on factors affecting Suprelorin(®) reversibility.
Topics: Animals; Animals, Wild; Animals, Zoo; Canidae; Contraceptive Agents; Drug Implants; Endangered Species; Felidae; Female; Megestrol Acetate; Practice Guidelines as Topic; Triptorelin Pamoate
PubMed: 23279543
DOI: 10.1111/rda.12004