Detection of doping peptides and basic drugs in equine urine using liquid chromatography-mass spectrometry.

The abuse of prohibited agents including peptides and basic small-molecule drugs is an area of great concern in horseracing due to their high potential to act as doping agents. These compound classes include agents such as growth hormone-releasing peptides, peptide analgesics, beta-2-adrenergic receptor agonists, and quaternary ammonium drugs that can be challenging to detect and regulate because of their chemical properties and potential rapid elimination following administration. The use of highly sensitive and selective analytical techniques such as liquid chromatography-mass spectrometry (LC-MS) is necessary to provide coverage of these substances and their potential metabolites. This study describes the development and validation of methodology capable of the detection of over 50 different peptide-based doping agents, related secretagogues, quaternary ammonium drugs, and other challenging small molecules in equine urine following solid-phase extraction using a mixed mode weak cation exchange sorbent. Following sample extraction, the compounds were analyzed using LC-MS with chromatographic separation via a reverse phase gradient and detection via selective reaction monitoring following introduction to a triple-stage quadrupole mass spectrometer using positive mode electrospray ionization. Validation parameters including limits of detection and quantitation, accuracy, precision, linear range, recovery, stability, and matrix effects were determined. Briefly, the limits of detection for most compounds were in the sub-ng/mL ranges with adequate precision and accuracy sufficient for an initial testing procedure. Stability studies indicated that most compounds were sufficiently stable to allow for effective screening using conditions commonly utilized in drug testing laboratories.

Leatherback sea turtle (Dermochelys coriacea) hatch success and essential and nonessential metals in eggs and embryos from nests in St. Kitts (2015).

Northwest Atlantic leatherback sea turtles (Dermochelys coriacea) are endangered and low hatch success limits potential for population recovery. We examined essential and nonessential metal concentrations in 43 eggs from nests on St. Kitts to determine if there was a relationship with hatch success. Whole homogenized embryos and undeveloped eggs contained detectable concentrations of arsenic, barium, copper, iron, selenium, vanadium, and zinc, but not beryllium, cadmium, chromium, cobalt, lead, mercury, molybdenum, and thallium. Of detected metals, only vanadium concentrations negatively correlated with hatch success (P = 0.01). Manganese and vanadium were associated with pneumonia occurring in the nest, and arsenic with renal mineralization. This study adds to the knowledge regarding baseline values for environmental contaminants in sea turtles, supporting the trend that leatherback eggs have relatively low concentrations of toxic metals, lacking a strong relationship with hatch success, and normally contain the essential elements copper, iron, selenium, and zinc.

Lead contamination in backyard chicken layer flocks in California.

Backyard layer chickens may be exposed to a variety of metals in the environment, including lead. The potential public health concerns associated with lead exposure prompted us to systematically screen liver samples from backyard layers submitted to the diagnostic laboratory to estimate the prevalence of lead exposure. Over a period of 1 y, we tested 1,476 chicken livers, of which 45 were found to have lead concentrations of 0.9-41 µg/g. The lead-positive cases were investigated by follow-up questions to the bird owners on the environment, general management of the flock, and egg consumption of family members. Lead concentrations in 14 pooled egg samples were determined, and a conservative estimate of daily exposure of family members to lead was made based on egg consumption. In some cases, estimated daily lead intake exceeded the recommended limits for lead consumption in children. Analysis of feed, water, and environmental samples did not identify a source of exposure in most cases. Only 34% of owners of lead-positive birds submitted eggs or environmental samples, indicating a lack of interest or financial concerns. In most cases, neither the case history nor postmortem findings were indicative of lead intoxication; without systematically testing all birds, some cases could have been missed. Our study highlights the need for backyard chicken owners, veterinarians, and public health personnel to be aware of the risk of lead exposure and undertake preventive and surveillance measures.

Interlaboratory trial for the measurement of total cobalt in equine urine and plasma by ICP-MS.

Cobalt is an essential mineral micronutrient and is regularly present in equine nutritional and feed supplements. Therefore, cobalt is naturally present at low concentrations in biological samples. The administration of cobalt chloride is considered to be blood doping and is thus prohibited. To control the misuse of cobalt, it was mandatory to establish an international threshold for cobalt in plasma and/or in urine. To achieve this goal, an international collaboration, consisting of an interlaboratory comparison between 5 laboratories for the urine study and 8 laboratories for the plasma study, has been undertaken. Quantification of cobalt in the biological samples was performed by inductively coupled plasma-mass spectrometry (ICP-MS). Ring tests were based on the analysis of 5 urine samples supplemented at concentrations ranging from 5 up to 500 ng/mL and 5 plasma samples spiked at concentrations ranging from 0.5 up to 25 ng/mL. The results obtained from the different laboratories were collected, compiled, and compared to assess the reproducibility and robustness of cobalt quantification measurements. The statistical approach for the ring test for total cobalt in urine was based on the determination of percentage deviations from the calculated means, while robust statistics based on the calculated median were applied to the ring test for total cobalt in plasma. The inter-laboratory comparisons in urine and in plasma were successful so that 97.6% of the urine samples and 97.5% of the plasma samples gave satisfactory results. Threshold values for cobalt in plasma and urine were established from data only obtained by laboratories involved in the ring test. Copyright © 2017 John Wiley & Sons, Ltd.