Authenticity analysis of oregano: development, validation and fitness for use of several food fingerprinting techniques.

Several analytical techniques, i.e. spectroscopic techniques as Near Infrared (NIR) and Mid-Infrared (MIR), Hyper Spectral Imaging (HSI), Gas Chromatography coupled to Mass Spectrometry (GC-MS) and Proton-transfer Reaction Time-of-Flight Mass spectrometry (PTR-TOF-MS), combined with chemometrics, are examined to evaluate their potential to solve different food authenticity questions on the case of oregano. In total, 102 oregano samples from one harvest season were analyzed for origin and variety assessment, 159 samples for adulteration-assessment and 72 samples for batch-to-batch control. The Gaussian Process Latent Variable Model (GP-LVM) was applied as technique to obtain a reduced two-dimensional space. A Random Forest Regression algorithm was used as regression model for the adulteration assessment. Prediction rates of more than 89% could be achieved for origin assessment. For variety assessment, prediction rates of more than 78% could be obtained. Batch-to-batch control could be successfully performed with NIR and PTR-TOF-MS. Detection of adulteration could be successfully performed from 10% on with HSI, NIR and PTR-TOF-MS.

Plasma serotonin in horses undergoing surgery for small intestinal colic.

This study compared serotonin concentrations in platelet poor plasma (PPP) from healthy horses and horses with surgical small intestinal (SI) colic, and evaluated their association with postoperative ileus, strangulation and non-survival. Plasma samples (with EDTA) from 33 horses with surgical SI colic were collected at several pre- and post-operative time points. Serotonin concentrations were determined using liquid-chromatography tandem mass spectrometry. Results were compared with those for 24 healthy control animals. The serotonin concentrations in PPP were significantly lower (P < 0.01) in pre- and post-operative samples from surgical SI colic horses compared to controls. However, no association with postoperative ileus or non-survival could be demonstrated at any time point. In this clinical study, plasma serotonin was not a suitable prognostic factor in horses with SI surgical colic.

Sérotonine plasmatique chez des chevaux subissant une chirurgie pour des coliques du petit intestin. Cette étude a comparé les concentrations de sérotonine dans le plasma faible en plaquettes (PFP) de chevaux en santé et de chevaux atteints de coliques chirurgicales du petit intestin et a évalué leur association avec l'occlusion intestinale postopératoire, la strangulation et la non-survie. Des échantillons de plasma (avec EDTA) ont été prélevés auprès de 33 chevaux atteints de coliques du petit intestin à plusieurs moments préopératoires et postopératoires. Les concentrations de sérotonine ont été déterminées à l'aide d'un spectromètre de masse LC-ESI-MS/MS. Les résultats ont été comparés avec ceux de 24 animaux témoins en santé. Les concentrations de sérotonine du PFP étaient significativement inférieures (P < 0,01) dans les échantillons préopératoires et postopératoires provenant des chevaux atteints de coliques du petit intestin comparativement aux animaux témoins. Cependant, aucune association avec l'occlusion intestinale postopératoire ou la non-survie n'a pu être démontrée à aucun moment. Dans cette étude clinique, la sérotonine plasmatique ne s'est pas avéré un facteur de pronostic approprié chez les chevaux atteints de coliques chirurgicales du petit intestin.(Traduit par Isabelle Vallières).

Quantification of 8-α-hydroxy-mutilin as marker residue for tiamulin in rabbit tissues by high-performance liquid chromatography-mass spectrometry.

For the first time, a sensitive and specific method was developed and fully validated for the quantification of the EU marker residue of tiamulin, 8-α-hydroxy-mutilin, in rabbit muscle and liver tissues using liquid chromatography combined with positive heated electrospray ionization triple quadrupole mass spectrometry. The mass spectrometer was operated in the selected reaction monitoring (SRM) mode with selection of the [M + H](+) ion in both quadrupoles 1 and 3, resulting in the SRM transition m/z 337.25 > 337.25 for quantification. Chromatography was performed using a Hypersil Gold C18 column using a gradient elution program with water and methanol as mobile phases. The sample preparation procedure for the analysis of 8-α-hydroxy-mutilin in liver and muscle samples consisted of three main steps: (1) extraction of the tissue matrix using 0.1 N hydrochloric acid/acetone (50/50, v/v), (2) hydrolysis of tiamulin and metabolites to 8-α-hydroxy-mutilin in alkaline medium at 45 °C, and (3) liquid-liquid extraction in acidic medium using ethyl acetate. This is the first method presenting fully validated results, encompassing a linearity of 50 to 2,000 µg/kg, within-run and between-run accuracy and precision, limit of quantification (50 µg/kg for both muscle and liver tissues), limit of detection (muscle, 11.9 µg/kg; liver, 20.6 µg/kg), extraction recovery (muscle, 66.2%; liver, 75.5%), signal suppression and enhancement (muscle, 51.7%; liver, 43.3%), carryover, applicability and practicability, and stability during storage and analysis. This novel method is therefore sensitive enough to be used for residue depletion studies of tiamulin in rabbits and for food safety monitoring with respect to MRL compliance of residues.

Comparative method validation for closantel determination in cattle and sheep milk according to European Union Volume 8 and Veterinary International Conference on Harmonization guidelines.

A specific and sensitive LC-MS/MS method was developed for quantitative determination of closantel in bovine and ovine colostrum and tank milk. Sample preparation consisted of extracting milk samples with acetonitrile/acetone (80/20, v/v) followed by SPE clean-up with Oasis mixed anion exchange columns. After elution with 5% formic acid in acetonitrile and evaporation to dryness, the residue was reconstituted in acetonitrile and water. HPLC separation was achieved on a Zorbax Eclipse Plus C18 column and a gradient elution program with 1mM ammonium acetate in water and acetonitrile. For closantel determination in bovine milk, the method was validated according to EU Volume 8 guidelines whereas for ovine milk both EU Volume 8 and VICH GL49 criteria were applied. The linear range of the method is between 10 and 2000 µg/kg, the limit of quantification 10 µg/kg and limit of detection is 0.63 and 0.32 µg/kg for sheep colostrum and tank milk and 1.27 and 1.24 µg/kg for cattle colostrum and tank milk, respectively. Both guidelines cover a similar set of parameters (linearity, accuracy, precision, limit of detection and limit of quantification), although the acceptance criteria might differ (accuracy and precision) or no specific acceptability ranges are specified in neither guidelines (LOD and LOQ). For some parameters, only one of the guidelines indicates acceptance criteria: EU Volume 8 for applicability, practicability and susceptibility and VICH GL 49 for linearity, specificity and analyte stability.

Comparative analysis of serotonin in equine plasma with liquid chromatography--tandem mass spectrometry and enzyme-linked immunosorbent assay.

Serotonin is regularly measured in equine platelet-poor plasma in research settings. However, reported reference values vary between studies, partially because plasma serotonin concentrations are very low and a reliable and affordable detection method is lacking. A simple, rapid, and sensitive method for serotonin determination in equine platelet-poor plasma using liquid chromatography--tandem mass spectrometry (LC-MS/MS) was developed and validated. Results of a commercially available enzyme-linked immunosorbent assay (ELISA) were compared to the LC-MS/MS results, in order to validate a test more suitable for use in a clinical situation. For LC-MS/MS, 500 µl of plasma was required, and deuterated serotonin was used as an internal standard. The sample preparation was based upon a simple liquid extraction into ethyl acetate. Chromatographic separation was performed with an acetic acid--acetonitrile mobile phase gradient elution. Linearity was demonstrated between 3 ng/ml and 100 ng/ml. A limit of quantification of 3 ng/ml was achieved, corresponding to a limit of detection of 0.10 ng/ml. Comparison of LC-MS/MS and ELISA with Passing-Bablok regression and Bland--Altman plotting showed a poor agreement between the 2 methods, with an increasing difference within the higher range of measurements. Caution is needed when extrapolating results from sources using different analytical techniques.

Evaluation of the pharmacokinetics and bioavailability of intravenously and orally administered amiodarone in horses.

OBJECTIVE: To determine the clinical effects and pharmacokinetics of amiodarone after single doses of 5 mg/kg administered orally or intravenously. ANIMALS: 6 healthy adult horses. PROCEDURE: In a cross over study, clinical signs and electrocardiographic variables were monitored and plasma and urine samples were collected. A liquid chromatography-mass spectrometry method was used to determine the percentage of protein binding and to measure plasma and urine concentrations of amiodarone and the active metabolite desethylamiodarone. RESULTS: No adverse clinical signs were observed. After IV administration, median terminal elimination half-lives of amiodarone and desethylamiodarone were 51.1 and 75.3 hours, respectively. Clearance was 0.35 L/kg x h, and the apparent volume of distribution for amiodarone was 31.1 L/kg. The peak plasma desethylamiodarone concentration of 0.08 microg/mL was attained 2.7 hours after IV administration. Neither parent drug nor metabolite was detected in urine, and protein binding of amiodarone was 96%. After oral administration of amiodarone, absorption of amiodarone was slow and variable; bioavailability ranged from 6.0% to 33.7%. The peak plasma amiodarone concentration of 0.14 microg/mL was attained 7.0 hours after oral administration and the peak plasma desethylamiodarone concentration of 0.03 microg/mL was attained 8.0 hours after administration. Median elimination half-lives of amiodarone and desethylamiodarone were 24.1 and 58.6 hours, respectively. CONCLUSION AND CLINICAL RELEVANCE: Results indicate that the pharmacokinetic distribution of amiodarone is multicompartmental. This information is useful for determining treatment regimens for horses with arrythmias. Amiodarone has low bioavailability after oral administration, does not undergo renal excretion, and is highly protein-bound in horses.