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The international production of farmed fish has been growing continuously over recent years. Until now few veterinary drugs have been approved by the European Union for use in aquaculture, and this has favored the off-label use of products authorized for use in food-producing animal species different from fishes among fish farmers. Adequate field studies are lacking, especially for those species called minor species which are consumed extensively only in some European countries. In the present investigation we studied the depletion of the fluoroquinolone antibacterial enrofloxacin over time in a minor species, the rainbow trout (Oncorhynchus mykiss), reared on a real fish farm and treated with medicated feed (10 mg kg of trout body weight ؊1 day ؊1 ). Edible tissue samples (muscle plus skin in natural proportions) and fish bone samples were analyzed for enrofloxacin and for its major metabolite, ciprofloxacin, by high-performance liquid chromatography with fluorescence detection at different times after the end of treatment. Our results show that at 500°C-day (in which degree-days are calculated by multiplying the mean daily water temperature by the total number of days on which the temperature was measured), which is the minimum withdrawal period established by European Economic Commission Directive No. 82/2001 for any type of product administered off-label, edible trout tissues might still contain about 170 g of enrofloxacin kg ؊1 , whereas the maximum residue level for enrofloxacin plus ciprofloxacin is set at 100 g kg ؊1 . To our knowledge, no studies of the depletion of enrofloxacin in rainbow trout have been performed. On the basis of the data obtained in the present study, we suggest a more appropriate withdrawal time of 816°C-day for the sum of enrofloxacin plus ciprofloxacin levels in rainbow trout muscle plus skin tissues.

In recent years, erythromycin has received considerable attention for its therapeutic efficacy against some bacterial kidney diseases in aquaculture and, therefore, suitable and sensitive analytical methods to monitor erythromycin residues in fish are required. A fast sample treatment followed by an LC-ESI-MS/MS method is described for the purification, identification, and quantification of erythromycin A residues in fish. After two extractions with acetonitrile, samples were defatted with n-hexane, filtered, and analyzed by tandem mass spectrometry. Three characteristic transition reactions (m/z 734 --> 716, 734 --> 576, and 734 --> 558) in multiple reaction monitoring were tested for the determination and confirmation of erythromycin A. The method was in-house validated through the determination of precision, accuracy, specificity, stability, calibration curve, decision limit (CCalpha), and detection capability (CCbeta), in accordance with European Commission Decision 657/2002. The coefficients of variation ranged from 1.8 to 9.4% and from 7.5 to 10.9% for intra- and interday repeatability, respectively. Recovery data were also satisfactory, with values varying from 85 to 97%. The method was specific, stable, and robust enough for the required purposes. The calibration curve showed a good linearity in the whole range of the tested concentrations (0-1000 microg kg(-1)) with a correlation coefficient (r2) equal to 0.9956. CCalpha and CCbeta were found to be 220 and 238 microg kg(-1), respectively.

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Aquaculture production has notably increased in the last decades, mainly thanks to intensive farming. Together with market globalization, this gives rise to the spreading of several fish diseases, thus increasing the demand for veterinary drugs for aquatic species. Nonetheless, very few chemicals are registered for use in aquaculture, and fish farmers are often forced to resort to off-label use of drugs authorized for other food-producing animal species. Rainbow trout is the major farmed fish species in Italy and the second one in Europe. Erythromycin is the antibiotic of choice against gram-positive cocci, the major concern for trout farming, but it is not yet registered for aquaculture use in most European countries. The aim of this study was to follow the depletion of erythromycin in rainbow trout (Oncorhynchus mykiss), after its administration at 100 mg kg ؊1 trout body weight day ؊1 for 21 days through medicated feed (water temperature, 11.5°C). Erythromycin residues in fish muscle plus skin in natural proportion were determined by a validated liquid chromatography-electrospray ionization-tandem mass spectrometry method. Interpolation of our data, following European Agency for the Evaluation of Medicinal Products guidelines, gives a withdrawal time of 255°C-days (°C-day ‫؍‬ water temperature ؋ days), thus showing that the general value (500°C-day) recommended by the Council Directive (EEC) no. 82/2001 for off-label drug use in aquaculture would be too conservative in this case, with excessive costs for the farmers. Our study provides preliminary data for a more prudent use of erythromycin in rainbow trout, suggesting a possible withdrawal time after treatment.

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