Short-term stability of testosterone and epitestosterone conjugates in urine samples: quantification by liquid chromatography-linear ion trap mass spectrometry.

A simple method using liquid chromatography-linear ion trap mass spectrometry for simultaneous determination of testosterone glucuronide (TG), testosterone sulfate (TS), epitestosterone glucuronide (EG) and epitestosterone sulfate (ES) in urine samples was developed. For validation purposes, a urine containing no detectable amount of TG, TS and EG was selected and fortified with steroid conjugate standards. Quantification was performed using deuterated testosterone conjugates to correct for ion suppression/enhancement during ESI. Assay validation was performed in terms of lower limit of detection (1-3ng/mL), recovery (89-101%), intraday precision (2.0-6.8%), interday precision (3.4-9.6%) and accuracy (101-103%). Application of the method to short-term stability testing of urine samples at temperature ranging from 4 to 37 degrees C during a time-storage of a week lead to the conclusion that addition of sodium azide (10mg/mL) is required for preservation of the analytes.

Measurement of β-tryptase in postmortem serum, pericardial fluid, urine and vitreous humor in the forensic setting.

In the realm of forensic pathology, β-tryptase measurement for diagnostic purposes is performed in postmortem serum obtained from femoral blood. This may be partially or completely unavailable in some specific cases, such as infant autopsies and severely damaged bodies. The aim of this study was to investigate the usefulness of determining β-tryptase levels for diagnostic purposes in alternative biological samples. Urine, vitreous humor and pericardial fluid were selected and measured in 94 subjects including: fatal anaphylaxis following contrast material administration (6 cases), hypothermia (10 cases), diabetic ketoacidosis (10 cases), gunshot suicide (10 cases), heroin injection-related deaths (18 cases), trauma (10 cases), sudden death with minimal coronary atherosclerosis (10 cases), severe coronary atherosclerosis without myocardial infarction (10 cases) and severe coronary atherosclerosis with myocardial infarction (10 cases). Postmortem serum and pericardial fluid β-tryptase levels higher than the clinical reference value (11.4ng/ml) were systematically identified in fatal anaphylaxis following contrast material administration and 6 cases unrelated to anaphylaxis. β-tryptase concentrations in urine and vitreous humor were lower than the clinical reference value in all cases included in this study. Determination of β-tryptase in pericardial fluid appears to be a possible alternative to postmortem serum in the early postmortem period when femoral blood cannot be collected during autopsy and biochemical investigations are required to objectify increased β-tryptase levels.

Effect of anionic salts on some blood and urine minerals, acid-base balance and udder oedema of dry pregnant cows

Selostus: Anionisten suolojen vaikutus ummessa olevien lehmien veren ja virtsan kivennäisiin, happo-emästasapainoon sekä utarepöhöön

Detection in urine of 4-methyl-2-hexaneamine, a doping agent.

Stimulants are banned in-competition for all categories of sports by the World Anti-Doping Agency. A simple liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay employing electrospray ionisation in positive mode was developed in that work for the quantification in urine specimens of 4-methyl-2-hexaneamine, a primary amine exhibiting sympathomimetic properties. Following a simple pretreatment procedure, the analyte was separated using a gradient mobile phase on reverse phase C8 column. Selected reaction monitoring m/z 116.2-->57.3 was specific for detection of 4-methyl-2-hexaneamine and the assay exhibited a linear dynamic range of 50-700 ng/mL. The validated method has been successfully applied to analyze the target compound in food supplements as well as in urine specimens. The administered drug (40 mg) was detected at the level of 350 ng/mL in the urine up to 4 days.

Fluorescence flow cytometer to determine urine particle reference intervals in doping control samples.

Background: Urine is still the matrix of choice to fight against doping, because it can be collected non-invasively during anti-doping tests. Most of the World Anti-Doping Agency's accredited laboratories have more than 20 years experience in analyzing this biological fluid and the majority of the compounds listed in the 2010 Prohibited List - International Standard are eliminated through the urinary apparatus. Storing and transporting urine samples for doping analyses does not include a specific protocol to prevent microbial and thermal degradation. The use of a rapid and reliable screening method could enable determine reference intervals for urine specimens in doping control samples and evaluate notably the prevalence of microbial contamination known to be responsible for the degradation of chemical substances in urine.Methods: The Sysmex(R) UF-500i is a recent urine flow cytometer analyzer capable of quantifying BACT and other urinary particles such as RBC, WBC, EC, DEBRIS, CAST, PATH. CAST, YLC, SRC as well as measuring urine conductivity. To determine urine anti-doping reference intervals, 501 samples received in our laboratory over a period of two months were submitted to an immediate examination. All samples were collected and then transported at room temperature. Analysis of variance was performed to test the effects of factors such as gender, test type [in-competition, out-of-competition] and delivery time.Results: The data obtained showed that most of the urine samples were highly contaminated with bacteria. The other urine particles were also very different according to the factors.Conclusions: The Sysmex(R) UF-500i was capable of providing a snapshot of urine particles present in the samples at the time of the delivery to the laboratory. These particles, BACT in particular, gave a good idea of the possible microbial degradation which had and/or could have occurred in the sample. This information could be used as the first quality control set up in WADA (World Anti-Doping Agency) accredited laboratories to determine if steroid profiles, endogenous and prohibited substances have possibly been altered. (C) 2011 Elsevier Ireland Ltd. All rights reserved.

Synthetic calibrators for the analysis of total metanephrines in urine: revisiting the conditions of hydrolysis.

BACKGROUND: The quantification of total (free+sulfated) metanephrines in urine is recommended to diagnose pheochromocytoma. Urinary metanephrines include metanephrine itself, normetanephrine and methoxytyramine, mainly in the form of sulfate conjugates (60-80%). Their determination requires the hydrolysis of the sulfate ester moiety to allow electrochemical oxidation of the phenolic group. Commercially available urine calibrators and controls contain essentially free, unhydrolysable metanephrines which are not representative of native urines. The lack of appropriate calibrators may lead to uncertainty regarding the completion of the hydrolysis of sulfated metanephrines, resulting in incorrect quantification. METHODS: We used chemically synthesized sulfated metanephrines to establish whether the procedure most frequently recommended for commercial kits (pH 1.0 for 30 min over a boiling water bath) ensures their complete hydrolysis. RESULTS: We found that sulfated metanephrines differ in their optimum pH to obtain complete hydrolysis. Highest yields and minimal variance were established for incubation at pH 0.7-0.9 during 20 min. CONCLUSION: Urinary pH should be carefully controlled to ensure an efficient and reproducible hydrolysis of sulfated metanephrines. Synthetic sulfated metanephrines represent the optimal material for calibrators and proficiency testing to improve inter-laboratory accuracy.

Prevalence of substance use in a Swiss psychiatric hospital: interview reports and urine screening.

BACKGROUND: Co-morbid substance misuse is common in psychiatric disorders, has potentially severe adverse consequences and may be frequently undetected. AIMS: To measure the prevalence of substance use among patients admitted to a Swiss psychiatric hospital and to examine the potential utility of routine urine drug screening in this setting. METHOD: 266 inpatients were included. 238 patients completed the interview and 240 underwent a urine drug screening. RESULTS: Lifetime prevalence of substance use among psychiatric patients was very high for alcohol (98%; 95% CI: 96-100), benzodiazepines (86%; 95% CI: 82-91) and cannabis (53%; 95% CI: 47-60), but also for "hard drugs" like cocaine (25% ; 95% CI: 19-30) or opiates (20%; 95% CI: 15-25). Regular current use of alcohol (32%; 95% CI: 26-38) or cannabis (17%; 95% CI: 12-22) was the most frequent. Substance use was associated with male sex, younger age, unmarried status and nicotine smoking. Urine screening confirms reports from patients on recent use, and remained positive for cannabis during hospitalisation, but not for cocaine nor for opiates. CONCLUSION: Substance use is frequent among psychiatric patients. Systematic interviewing of patients about their substance use remains essential, and is usually confirmed by urine screening. Urine screening can be useful to provide specific answers about recent use.

In Vivo Profiling and Distribution of Known and Novel Phase I and Phase II Metabolites of Efavirenz in Plasma, Urine, and Cerebrospinal Fluid.

Efavirenz (EFV) is principally metabolized by CYP2B6 to 8-hydroxy-efavirenz (8OH-EFV) and to a lesser extent by CYP2A6 to 7-hydroxy-efavirenz (7OH-EFV). So far, most metabolite profile analyses have been restricted to 8OH-EFV, 7OH-EFV, and EFV-N-glucuronide, even though these metabolites represent a minor percentage of EFV metabolites present in vivo. We have performed a quantitative phase I and II metabolite profile analysis by tandem mass spectrometry of plasma, cerebrospinal fluid (CSF), and urine samples in 71 human immunodeficiency virus patients taking efavirenz, prior to and after enzymatic (glucuronidase and sulfatase) hydrolysis. We have shown that phase II metabolites constitute the major part of the known circulating efavirenz species in humans. The 8OH-EFV-glucuronide (gln) and 8OH-EFV-sulfate (identified for the first time) in humans were found to be 64- and 7-fold higher than the parent 8OH-EFV, respectively. In individuals (n = 67) genotyped for CYP2B6, 2A6, and CYP3A metabolic pathways, 8OH-EFV/EFV ratios in plasma were an index of CYP2B6 phenotypic activity (P < 0.0001), which was also reflected by phase II metabolites 8OH-EFV-glucuronide/EFV and 8OH-EFV-sulfate/EFV ratios. Neither EFV nor 8OH-EFV, nor any other considered metabolites in plasma were associated with an increased risk of central nervous system (CNS) toxicity. In CSF, 8OH-EFV levels were not influenced by CYP2B6 genotypes and did not predict CNS toxicity. The phase II metabolites 8OH-EFV-gln, 8OH-EFV-sulfate, and 7OH-EFV-gln were present in CSF at 2- to 9-fold higher concentrations than 8OH-EFV. The potential contribution of known and previously unreported EFV metabolites in CSF to the neuropsychological effects of efavirenz needs to be further examined in larger cohort studies.

Blood pressure reductions following catheter-based renal denervation are not related to improvements in adherence to antihypertensive drugs measured by urine/plasma toxicological analysis.

Renal denervation can reduce blood pressure in patients with uncontrolled hypertension. The adherence to prescribed antihypertensive medication following renal denervation is unknown. This study investigated adherence to prescribed antihypertensive treatment by liquid chromatography-high resolution tandem mass spectrometry in plasma and urine at baseline and 6 months after renal denervation in 100 patients with resistant hypertension, defined as baseline office systolic blood pressure ≥140 mmHg despite treatment with ≥3 antihypertensive agents. At baseline, complete adherence to all prescribed antihypertensive agents was observed in 52 patients, 46 patients were partially adherent, and two patients were completely non-adherent. Baseline office blood pressure was 167/88 ± 19/16 mmHg with a corresponding 24-h blood pressure of 154/86 ± 15/13 mmHg. Renal denervation significantly reduced office and ambulatory blood pressure at 6-month follow-up by 15/5 mmHg (p < 0.001/p < 0.001) and 8/4 mmHg (p < 0.001/p = 0.001), respectively. Mean adherence to prescribed treatment was significantly reduced from 85.0 % at baseline to 80.7 %, 6 months after renal denervation (p = 0.005). The blood pressure decrease was not explained by improvements in adherence following the procedure. Patients not responding to treatment significantly reduced their drug intake following the procedure. Adherence was highest for angiotensin-converting enzyme inhibitors/angiotensin receptor blockers and beta blockers (>90 %) and lowest for vasodilators (21 %). In conclusion, renal denervation can reduce office and ambulatory blood pressure in patients with resistant hypertension despite a significant reduction in adherence to antihypertensive treatment after 6 months.