Antibiotics are active substances frequently used to treat and prevent diseases in animal husbandry, especially in swine and poultry farms. The use of manure as a fertilizer may lead to the dispersion of antibiotic residue into the environment and consequently the development of antibiotic-resistant bacteria. Most pharmaceutical active ingredients are excreted after administration, in some cases up to 90% of the consumed dose can be found in the feces and/or urine as parent compound. Therefore, due to antibiotic metabolism their residues can be easily detected in manure. This article describes a method for simultaneous analysis of ciprofloxacin, chlortetracycline, doxycycline, enrofloxacin, lincomycin, oxytetracycline, tetracycline, tiamulin, trimethoprim and tylosin in feces, liquid manure and digestate. Antibiotics were extracted from the different matrices with McIlvaine-Na2EDTA buffer solution and the extract was purified by the use two techniques: d-SPE and SPE (Strata-X-CW cartridges) and final eluent was analyzed by LC-MS and LC-MS/MS. The European Commission Decision 2002/657/EC was followed to conduct the validation of the method. Recoveries obtained from spiked pig and poultry feces and liquid manures samples ranged from 63% to 93% depending on analytes. The analysis of 70 samples (feces, liquid manure and digestate) revealed that 18 samples were positive for the presence of doxycycline, oxytetracycline, tetracycline, chlortetracycline, enrofloxacin, tiamulin and lincomycin. The results obtained in the presented study demonstrated that animal feces can be used as a non-invasive method detection antibiotic usage in animal production.

A new multi-compound method for the analysis of veterinary drugs, namely tiamulin, trimethoprim, tylosin, sulfadiazine and sulfamethazine was developed and validated in medicated feeds. After extraction, the samples were centrifuged, diluted in Milli-Q water, filtered and analysed by high performance liquid chromatography coupled to tandem mass spectrometry. The separation of the analytes was performed on a biphenyl column with a gradient of 0.1% formic acid in acetonitrile and 0.1% formic acid in Milli-Q water. Quantitative validation was done in accordance with the guidelines laid down in European Commission Decision 2002/657/EC. Method performances were evaluated by the following parameters: linearity (R < 0.99), precision (repeatability <14% and within-laboratory reproducibility <24%), recovery (73.58-115.21%), sensitivity, limit of detection (LOD), limit of quantification (LOQ), selectivity and expanded measurement uncertainty (k. = 2). The validated method was successfully applied to the 2 medicated feeds obtained from the interlaboratory studies and feed manufactures from Spain in August 2017. In these samples, tiamulin, tylosin and sulfamethazine were detected at the concentration levels declared by the manufacturers. The developed method can therefore be successfully used to routinely control the content and homogeneity of these antibacterial substances in medicated feed. Abbreviations AAFCO - Association of American Feed Control Officials; TYL - tylosin; TIAM - tiamulin fumarate; TRIM - trimethoprim; SDZ - sulfadiazine; SMZ - sulfamethazine; UV - ultraviolet detector; FLD - fluorescence detector; HPLC - high performance liquid chromatography; MS/MS - tandem mass spectrometry; LOD - limit of detection; LOQ - limit of quantification; CV - coefficient of variation; SD - standard deviation; U - uncertainty.

The use of medicated feed is a common practice in animal food production to improve animal health. Tetracyclines and β-Lactams are the groups that are most frequently added to this type of feed. The measurement of the concentration of the analytes in these types of samples is sometimes due to the matrix characteristic, and manufacturers are demanding fast, precise and reproducible methods. A rapid confirmatory method based on a simple extraction protocol using acidified methanol and followed by high performance liquid chromatography coupled to a tandem mass spectrometer for the quantification of four tetracyclines in feed is presented. Validation was performed following the guidelines of Decision 2002/657/EC. Results indicated that the four tetracyclines can be identified and quantified in a concentration range between 50 and 500 mg/kg with recoveries between 84% and 109% and RSD for precision under reproducible conditions between 12% and 16%. Satisfactory results were also obtained with interlaboratory studies and by comparing the method with an HPLC-Fluorescent method.

A confirmatory HPLC-MS/MS method for the determination of residues of 11 antibacterial substances from different therapeutic class (β-lactams, lincosamides, fluoroquinolones, macrolides, pleuromutilins and sulfonamides) in animal feeds has been developed. The sample preparation is based on extraction with 0.1% formic acid in acetonitrile. Separation of the analytes was performed on biphenyl column with a gradient of 0.1% formic acid in acetonitrile and 0.1% formic acid in Milli-Q water. The developed method was validated following the guidelines included in the European Union Commission Decision 2002/657/EC. Limits of detection ranging from 79.22 to 193.60 µg/kg; instrumental and analytical linearity coefficients were above 0.99 for matrix-match calibration; and relative recoveries ranging from 76.04% to 117.39%. Repeatability of the method was in the range of 2.41-19.76% (CV, %), whereas reproducibility ranged from 6.52 to 28.40% (CV %). The method shown to be efficient and precise for quantification of the 11 antibacterial substances in animal feed. The results demonstrate the feasibility of the method for routine use to monitor these substances in feed. The validated method was successfully applied to eight suspect feed samples collected from the Association of American Feed Control Officials (AFFCO) and feed manufactures from Galicia (Spain) in June and July 2017. Of these 8 non-target feeds, 5 were positive for the presence of tiamulin, tylosin and sulfadiazine.

Taking into consideration the maximum level (ML) for coccidiostats included in the European Regulation 574/2011 and the fact that the presence of residues of sulphonamides in non-target feed is forbidden, the aim of this article is to present an analytical method based on HPLC-MS/MS for the identification and quantification of sulphonamides and coccidiostats in non-target feeds. The method was validated following Commission Decision 2002/657/EC, and recovery, repeatability and reproducibility were within the limits established in the Decision. For coccidiostats, the decision limit and detection capability were calculated for the different species taking into account the ML allowed in Regulation 574/2011. The applicability of the method was investigated in 50 feed samples collected from dairy farms, 50 obtained from feed mills and 10 interlaboratory feed samples.