Development and validation of a gas chromatography/mass spectrometry procedure for confirmation of para-toluenesulfonamide in edible fish fillet tissue.

Chloramine-T is a disinfectant being developed as a treatment for bacterial gill disease in cultured fish. As part of the drug approval process, a method is required for the confirmation of chloramine-T residues in edible fish tissue. The marker residue that will be used to determine the depletion of chloramine-T residues from the edible tissue of treated fish is para-toluenesulfonamide (p-TSA), a metabolite of chloramine-T. The development and validation of a procedure for the confirmation of p-TSA is described. Homogenized fish tissue is dried by mixing with anhydrous sodium sulfate, and the mixture is extracted with methylene chloride. The extract is passed through a silica gel solid-phase extraction column, from which p-TSA is subsequently eluted with acetonitrile. The acetonitrile extract is evaporated, and the oily residue is dissolved in hexane. The hexane solution is shaken with fresh acetonitrile. The acetonitrile solution is evaporated and the residue is redissolved in dilute potassium hydroxide solution. The aqueous solution is extracted with methylene chloride to further remove more of the fat co-extractive. The aqueous solution is reacted with pentafluorobenzyl bromide in presence of tetrabutylammonium hydrogensulfate. The resulting di-(pentafluorobenzyl) derivative of p-TSA is analyzed by gas chromatography/mass spectrometry. This method permits the confirmation of p-TSA in edible fish tissue at 20 ppb.

Simple, rapid determination of enrofloxacin and ciprofloxacin in bovine milk and plasma by high-performance liquid chromatography with fluorescence detection.

A rapid and simple procedure for determination of enrofloxacin and ciprofloxacin in bovine milk and plasma is described. Protein precipitation from both milk and plasma samples was achieved by addition of acetonitrile and phosphoric acid. Acetonitrile was removed with methylene chloride, leaving enrofloxacin and ciprofloxacin in the acidic aqueous extract. The aqueous extract was analyzed by high-performance liquid chromatography (HPLC) with fluorescence detection. The limit of quantitation (LOQ) for enrofloxacin and ciprofloxacin in milk was found to be 2ng/ml. LOQ for enrofloxacin and ciprofloxacin in plasma was found to be 1ng/ml. Linear calibration curves were obtained with correlation coefficient (r(2)) >/=0.99. Analysis of quality control (QC) samples gave results within +/-10% of the nominal values. Inter-assay precision for the analysis of milk QC samples were in the ranges: 4.63-12.49% (for enrofloxacin) and 4.67-9.86% (for ciprofloxacin). Inter-assay precision for the analysis of plasma QC samples were in the ranges: 6.60-17.31% (for enrofloxacin) and 6.14-13.87% (for ciprofloxacin). Intra-assay precision for the analysis of milk QC samples were in the following ranges: 3.65-7.21% (for enrofloxacin) and 1.58-14.28% (for ciprofloxacin). Intra-assay precision for the analysis of plasma QC samples were in the following ranges: 2.17-16.95% (for enrofloxacin) and 3.31-16.31% (for ciprofloxacin). The effectiveness of protein precipitants other than phosphoric acid was investigated. The method described has been applied to a study of the pharmacokinetics of enrofloxacin and ciprofloxacin in lactating dairy cows and beef steers.