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Abstract: The Commission Decision 2002/657/EC is a fundamental reference document for the UE laboratories involved in residue analysis although its implementation has caused some difficulties in the requirements interpretation. In this work a pragmatic validation approach of a quantitative confirmatory method for the detection of 17-alpha-(α-NT) and 17-beta-19-nortestosterone (β-NT) in bovine urine by gas chromatography mass spectrometry is proposed. The 19-nortestosterone is a banned anabolic steroid for which no minimum required performance limit (MRPL) has been laid down, therefore the limit reported in Italian Residue Monitoring Plan (2μgL−1) has been considered the reference level to evaluate the method performances. The decision limit (CCα) and the detection capability (CCβ) were obtained by the calibration curve procedure. The minimum required performance level (mrpl), which represents the starting concentration of the calibration curves, was preliminary fixed estimating the results dispersion of blank urine samples fortified at 2μgL−1 for each isomer. The found CCα and CCβ were 1.5 and 1.9μgL−1 for α-NT and 1.2 and 1.4μgL−1 for β-NT. The precision (repeatability and within-laboratory reproducibility) and recoveries were suitable for the investigated concentration range (1–3μgL−1). Finally, the method ruggedness (minor and major changes) has been also demonstrated. [Copyright &y& Elsevier]

A simple multi-residue method is described for assaying 13 quinolones (QNs; pipemidic acid, rufloxacin, enoxacin, ofloxacin, norfloxacin, ciprofloxacin, danofloxacin, enrofloxacin, difloxacin, cinoxacin, oxolinic acid, nalidixic acid and flumequine) in feeds. The samples were extracted by a metaphosphoric acid/acetonitrile mixture at pH 2.6 (ASE 200) and automatically purified onto OASIS HLB cartridges (ASPEC XL). The determination was achieved by liquid chromatography (LC) using a C5 analytical column (150×4.6 mm, 5 μm) either with photodiode-array or fluorescence detection. The procedure was validated by spiking a feed sample at three different levels (5, 10 and 25 mg kg−1) and linearity, detection limit, quantification limit, accuracy and precision were checked. The automated preparation of the sample permits a very fast analysis which is an important goal for routine purposes. [Copyright &y& Elsevier]

A novel method for the simultaneous identification and quantification of twelve aminoglycosides (AGs) and two colistins in meat and bovine milk has been developed. The analysis was carried out using liquid chromatography coupled to quadrupole-Orbitrap mass spectrometry (LC-Q-Orbitrap). Among the HILIC (Hydrophilic Interaction Liquid Chromatography) stationary phases tested, the bare silica Poroshell 120 provided the best results. The samples were extracted with an aqueous solution followed by an SPE clean up based on the weak cation exchange mechanism. The validation study was performed carrying out 72 experiments per matrix at six different concentrations in a range encompassing the Maximum Residue Limits. The recoveries were from 72 to 87% in meat (except colistins) and from 82 to 96% in milk. Repeatabilities and intra-lab reproducibilities were lower than 10 and 15%, respectively. Limits of detection were lower than or equal to 33 µg kg −1 . Finally, test materials containing AGs prepared for interlaboratory studies were successfully analysed. [ABSTRACT FROM AUTHOR]

The expression of CYP4F2, a form of cytochrome P-450 with proposed role in α-tocopherol and long-chain fatty acid metabolism, was explored in HepG2 and HepaRG human hepatocytes during ethanol toxicity. Cytotoxicity, ROS production, and JNK and ERK1/2 kinase signaling increased in a dose and time-dependent manner during ethanol treatments; CYP4F2 gene expression decreased, while other CYP4F forms, namely 4F11 and 12, increased along with 3A4 and 2E1 isoforms. α-Tocopherol antagonized the cytotoxicity and CYP4F2 gene repression effect of ethanol in HepG2 cells. Ethanol stimulated the tocopherol-ω-hydroxylase activity and the other steps of vitamin E metabolism, which points to a minor role of CYP4F2 in this metabolism of human hepatocytes. PPAR-γ and SREBP-1c followed the same expression pattern of CYP4F2 in response to ethanol and α-tocopherol treatments. Moreover, the pharmacological inhibition of PPAR-γ synergized with ethanol in decreasing CYP4F2 protein expression, which suggests a role of this nuclear receptor in CYP4F2 transcriptional regulation. In conclusion, ethanol toxicity modifies the CYP expression pattern of human hepatic cells impairing CYP4F2 transcription and protein expression. These changes were associated with a lowered expression of the fatty acid biosynthesis regulators PPAR-γ and SREBP-1c, and with an increased enzymatic catabolism of vitamin E. CYP4F2 gene repression and a sustained vitamin E metabolism appear to be independent effects of ethanol toxicity in human hepatocytes. [ABSTRACT FROM AUTHOR]

The aim of this study was to evaluate the safety of intra-articular (IA) lidocaine plus adrenaline for improving peri-operative analgesia in anaesthetized dogs undergoing arthroscopy of the elbow. A solution of lidocaine (L) 1.98% plus adrenaline 1:100.000 was administered via the IA route and its safety evaluated in terms of cardio-, neuro-, and chondro-toxicity. No bradycardia or hypotension was recorded from induction to the last observational time point. Signs of toxicity of the nervous system could have been masked by the general anaesthesia but lidocaine concentrations detected in the blood were lower than those thought to be capable of producing toxicity. The assessment of in vitro chondrotoxicity showed a dose- and time-dependent effect of lidocaine on the viability of articular cells. Adrenaline appeared to reduce the chondrotoxicity of 1% lidocaine, following an exposure of up to 30 min. [ABSTRACT FROM AUTHOR]

Abstract: A commercial enzyme-linked immunosorbent assay (ELISA) test has been applied for the qualitative screening analysis of chloramphenicol (CAP) in food of animal origin (muscle, eggs, milk and honey) at level corresponding to the European Union (EU) minimum required performance limit (MRPL) set for CAP analysis. Different analytical protocols were developed for the extraction and purification of CAP from the matrices under investigation. The ELISA kit validation study was performed according to the Commission Decision 2002/657/EC criteria established for qualitative screening methods. In this regard, the following parameters were determined: detection capability (CCβ), specificity/selectivity, applicability/ruggedness/stability. The CCβ value resulted below the MRPL (0.3μgkg-1) for all the examined matrices. No relevant interferences from matrix effects or structurally related substances (florfenicol (FF) and thiamphenicol (THP)) were observed. Slight variations of some critical factors in the sample pre-treatment and clean up for muscle were deliberately introduced for ruggedness evaluation and they did not result in any negative effect on the CAP detection. In addition, the recovery rates and precision at three concentration levels (0.30–0.45–0.60μgkg-1) were evaluated for all matrices under investigation. The proposed method is suitable for qualitative screening analysis of CAP in the above-mentioned food in conformity with the current EU performance requirements. [Copyright &y& Elsevier]