High-performance liquid chromatographic method with fluorescence detection for the screening and quantification of oxolinic acid, flumequine and sarafloxacin in fish.

A previously published liquid chromatographic method for determining residues of nine quinolones in chicken, porcine, bovine and ovine muscle was adapted and applied to fish tissue for simultaneous determination of three quinolones (flumequine, oxolinic acid and sarafloxacin). The analytes were extracted from homogenised muscle using an acetonitrile basic solution. After centrifugation, partial evaporation and cleaning with hexane, direct injection was possible. Separation was achieved on PLRP-S column and detection was performed with a programmable fluorescence detector. Chromatographic conditions were optimised to be compatible with the determination of the three quinolones in a single run. The linearity, recovery, accuracy and precision of the method were evaluated from fortified tissue samples at concentration levels ranging from 15 to 120 microg kg(-1) for sarafloxacin and 75 to 600 microg kg(-1) for oxolinic acid and flumequine according to the EU maximum residue limit of each quinolone. The limits of detection were estimated to be 2, 5 and 7 microg kg(-1), respectively, for sarafloxacin, oxolinic acid and flumequine. The limits of quantification were validated at 15 microg kg(-1) for sarafloxacin and 75 microg kg(-1) for oxolinic acid and flumequine. Mean extraction recoveries of quinolones in fish ranged from 56.9 to 71.0%. This simple and rapid method is suitable for residue control.

Quantitation of nine quinolones in chicken tissues by high-performance liquid chromatography with fluorescence detection.

A simple reversed-phase high-performance liquid chromatographic method was developed and validated for simultaneous analysis of nine quinolones (ciprofloxacin, danofloxacin, difloxacin, enrofloxacin, flumequine, marbofloxacin, nalidixic acid, oxolinic acid, sarafloxacin) in chicken tissue. The analytes were extracted from homogenized muscle using an acetonitrile basic solution. After centrifugation and partial evaporation, direct injection was possible. Three different HPLC conditions were applied to quantify the residual quinolones. Separation was achieved on a PLRP-S column and detection was performed with a monochromator fluorescence detector. The recovery, the limit of detection, the limit of quantification, the accuracy and the precision of the method were evaluated from spiked tissue samples at concentration levels ranging from 15 microg kg(-1) to 300 microg kg(-1) according to the maximum residue limit of each quinolone. This method is also suitable for porcine, bovine, ovine and fish muscle tissue.

6-[18F]fluoro-L-dopa uptake and [76Br]bromolisuride binding in the excitotoxically lesioned caudate-putamen of nonhuman primates studied using positron emission tomography.

The functional status of the dopaminergic system following striatal excitotoxic lesions was studied in living baboons by positron emission tomography (PET) using 6-[18F]fluoro-L-dopa as specific tracer for the presynaptic dopaminergic terminals and [76Br]bromolisuride as selective dopamine D2-receptor marker. The glutamate receptor agonist ibotenic acid (IA) was injected into the right caudate-putamen of six baboons to induce a neuropathological and behavioral model of Huntington's disease (HD). In vivo PET studies performed 3 to 6 months after the IA injections showed that subtotal excitotoxic lesions of the CP were accompanied by changes in the kinetic of [76Br]bromolisuride binding indicating a dose-dependent reduction in binding sites in the lesioned striatum of all IA-injected animals. In the most severely lesioned animals, there was also a decrease in the uptake of the nigrostriatal dopaminergic marker. The loss of D2-receptors and decrease in striatal dopamine uptake are consistent with clinical and postmortem findings in HD. In addition, the decrease in 6-[18F]fluoro-L-dopa uptake confirms previous studies performed in a rat model of HD suggesting a continuous decline of nigral dopamine cell function following destruction of their intrinsic striatal target neurons. The results of our experience to date in PET studies of 6-[18F]fluoro-L-dopa and [76Br]bromolisuride binding in IA-lesioned primates indicate that PET can identify effects of cell loss on markers of pre- and postsynaptic function in the striatum of living subjects.

Quantitative evaluation of benzodiazepine receptors in live Papio papio baboons using positron emission tomography.

The binding of the 11C-labeled benzodiazepine antagonist Ro 15-1788 (flumazenil) was measured in the neocortex of live Papio papio baboons by positron emission tomography. This allowed us to calculate in vivo (i.e., at physiological temperature, neurotransmitters concentrations, and ionic environment) the apparent density of available benzodiazepine receptors (B'max) and the dissociation constant of Ro 15-1788 (Kd). By coadministering increasing doses of unlabeled Ro 15-1788 with [11C]Ro 15-1788 and assuming that nonsaturable radioactivity indicated the free ligand concentration, we were able to obtain saturation isotherms. We showed that a state of quasiequilibrium was reached 50 min after the administration of the radioligand. Linear Scatchard plots allowed us to calculate B'max at 78 and 50 pmol/ml of cerebral tissue in the occipital and frontal cortices, respectively. In both these areas, Kd is on the order of 6 nM, with a Hill number very close to unity. This indicates that Ro 15-1788 binds in vivo with high affinity to an homogeneous population of saturable sites. A similar measurement was carried out on a naturally photosensitive P. papio baboon. Absolute values of B'max, Kd, and Hill number were similar to those of the control baboons. Although results concerning this baboon can only be considered as a case report, this similarity may suggest that its epileptic syndrome is not related to a large change in B'max or Kd, at least in occipital and frontal cortices. Our results showed that quantitative estimation by positron emission tomography of some characteristics of benzodiazepine receptors is possible in live baboons and may represent a supplementary tool for investigating further the molecular mechanisms of benzodiazepine receptor function in physiological and physiopathological conditions. We suggest that a similar method of quantification of classic in vivo [3H]Ro 15-1788 binding could be usefully adapted when studying rodent models of epilepsy, stress, and other neuropsychological disorders. On the other hand, the similarity between the B'max and Kd values we obtained in baboons and those recently reported in humans using similar methods emphasizes that most of the in vivo characteristics of the benzodiazepine receptors of baboons are very close to those of human benzodiazepine receptors. This confirms that P. papio baboons are a suitable animal model for studying the pharmacology of benzodiazepine receptor ligands before clinical applications in humans.