Abstract: Nalidixic acid and norfloxacin were determined in this work for the first time by applying kinetic methodology to complex formation reactions involving energy-transfer processes. By using initial-rate measurements, a fluorescence-based method based on the formation of the complexes of these compounds with terbium(III) in the presence of Tris buffer was developed. The formation rate of these complexes is very high, so the stopped-flow mixing technique is required in order to make kinetic measurements, which, in addition, allows the method to be automated. A second ligand, EDTA, is also used to decrease the reaction rate of both systems via ligand-exchange, which is necessary in order to obtain appropriate kinetic data. The reaction rate is obtained within only 0.1-0.2 s after the reactants are mixed, which allows ready application of the proposed method to routine analyzes. The calibration graph is linear over the range 0.02-7.0 µg mL-1 nalidixic acid and 0.4-9.0 µg mL-1 norfloxacin. The detection limit for nalidixic acid and norfloxacin is 0.006 and 0.13 µg mL-1, respectively. The selectivity and precision of the method were also studied. The proposed kinetic method was satisfactorily applied to the direct determination of these compounds in serum samples.

Abstract: A liquid-chromatographic (LC) method is described for determining residues of flumequine (FLU) and nalidixic (NAL), oxolinic (OXO), and piromidic (PIR) acids in catfish muscle. The identities of 3 of these residues are confirmed by gas chromatography-mass spectrometry (GC-MS). The extraction and cleanup procedures are the same for both determination and identification. Analyte isolation involves homogenizing the tissue with acetone, defatting the acetone extract with hexane, and extracting the compounds into chloroform. The extract is further purified by first partitioning into base and subsequently back-extracting into chloroform after acidifying the aqueous phase. After the solvent is evaporated, the residue is dissolved in mobile phase, and the analytes are determined by LC with fluorescence detection, excitation at 325 nm and emission at 365 nm. Catfish muscle was fortified with each quinolone at 5, 10, 20, 40, and 80 ng/g. Recoveries were 83-94%, with relative standard deviations (RSDs) of 5-7%. The method was evaluated also by a second analyst, who determined 4 quinolones added in combination. Average recoveries of quinolones from catfish fortified at 5, 10, and 20 ng/g were 78-90%, with RSDs of 3-6%. The presence in catfish muscle of incurred OXO, FLU, and NAL at the 10 ng/g level was confirmed by analyzing the decarboxylated quinolones by GC-MS. The relative abundances of all 5 major ions for OXO, FLU, and NAL were within 10% of those observed in spectra of standard compounds decarboxylated by the same method.

Abstract: Bleomycin A2 (I) and nalidixic acid (II) could be detected down to limits of 3 µM and 0.4 µM, respectively, by luminescence detection at 545 nm after excitation with Tb(III) at 318 nm (I) or 310 nm (II). For reversed-phase HPLC separation of I a column (15 cm x 4.6 mm i.d.) of Beckman ODS (5 µm) was used or for anion-exchange separation a column (25 cm x 4.1 mm i.d.) of Alltech Anion R (10 µm) was used with a Wescan guard cartridge. For measurement of I a mobile phase of 10 mM TbCl3 and 5 mM heptanesulfonic acid in aqueous 0.5% acetic acid of pH 6.8/methanol (11:9) was used. Linear response was shown over two orders of magnitude. For II the detection limit was established by a flow injection method and a suitable HPLC method for determination has still to be found.