Introduction: The article presents a rapid and simple analytical procedure for determination of four tetracyclines (oxytetracycline, tetracycline, chlortetracycline, and doxycycline) in animal medicated feedingstuffs. Material and Methods: Two-gramme samples were extracted by a Na2EDTA-McIlvaine buffer (pH 4)/methanol mixtures (40/60, v/v). The determination was achieved by liquid chromatography using a Zorbax Eclipse XDB C18 analytical column with mass spectrometer detection (LC-MS). Results: Recoveries of the antibiotics from spiked feed samples ranged from 78.2% to 113.5%. The LOD and LOQ for tetracyclines in feeds ranged from 2.8 to 4.2 and from 4.3 to 5.7 mg/kg, respectively. Conclusion: The method was successfully validated and proved to be efficient, precise, and useful for quantification of tetracyclines in medicated feedingstuffs.
A high-performance liquid chromatographic–diode array detector (HPLC-DAD) method for the determination of amoxicillin in medicated feedingstuffs was developed and validated. The method was used to investigate the quality requirements of animal feedingstuffs (declared content of active substance and feed homogeneity).
Material and Methods
Two-gram samples were extracted by potassium phosphate buffer solution. Extracts were filtered and directly analysed by HPLC-DAD without further clean-up. Amoxicillin was separated by acetonitrile and 0.01M phosphate buffer (pH 5.0) on a Phenomenex Luna C18 column.
This method provided average recoveries of 76.1 to 81.6% with coefficients of variation (CV, %) for repeatability and reproducibility in the ranges of 3.7–7.2% and 5.3–7.6%, respectively. The limit of detection was 51.2 mg/kg and limit of quantification was 103.0 mg/kg.
The method was successfully validated and proved to be efficient, precise, and useful for quantification of amoxicillin in medicated feedingstuffs.
A chromatographic procedure for determination of oxytetracycline (OXT), tetracycline (TC), chlorotetracycline (CTC), and doxycycline (DC) in water samples was developed and was applied for the analysis of water samples collected from poultry and pig farms and environmental water samples. Samples were acidified with trifluoroacetetic acid to pH 3 and further purified by solid phase extraction using Oasis HLB cartridges. The samples were dried up and redissolved in the mixture of oxalic acid and methanol. Separation was performed on reserved phase column (Phenomenex column C18 , 250 mm × 4.6 mm, 5 μm) by multistep gradient elution, and detection was carried out at 360 nm for OTC and TC, 370 nm for CTC, and 350 nm for DC. The tetracyclines were eluted with the mobile phase of 0.05 M oxalic acid (pH 2.5), acetonitrile, and methanol. This method provided average recoveries of 83.53% to 108.59%, with coefficient of variations (CVs) of 2.41% to 8.64% in the range of 10 to 1000 μg/L OTC, TC, CTC, and DC in water. The linearity for the tetracyclines was determined by HPLC-DAD in the range 10 to 1000 μg/L, with the correlation coefficient (R) > 0.99. The LOD and LOQ for the tetracyclines in water samples ranged from 1.51 to 4.00 and 2.51 to 5.93 μg/L, respectively.
High performance liquid chromatography method with diode array detection (HPLC-DAD) was developed and optimised for the determination of tetracyclines (TCs) in medicated feedingstuffs. The extraction of the analyte from feedingstuffs was performed with Na2EDTA-McIlvaine buffer (pH 2.5 and pH 4). The extracts were cleaned up by solid phase extraction using octadecyl cartridges (C18). The samples were dried up and redissolved in the mixture of oxalic acid and methanol. Separation was performed on reserved phase column (Phenomenex C18, 250 x 4.6 mm, 5 μm) by multistep gradient elution, which provided better chromatographic separation. The analysis was performed at a wavelength of 390 nm. Validation study of the method revealed that all obtained calibration curves showed good linearity R= 0.9985 for doxycycline (DC) and R= 0.9981 for chlorotetracycline (CTC) over the range of 25-2,000 mg/kg. The analytical procedure was successfully adapted for quantitative determination of DC and CTC in medicated feedingstuff samples. Validation included determination of specificity, linearity, and repeatability. Mean recovery for spiked samples was 93.1% for CTC and 93.2% for DC. The results of validation of the analytical procedure proved that presented method is efficient, precise and useful for quantification of DC and CTC in medicated feedingstuffs.
The paper describes a microbiological method for the detection of antibacterial substances in feedingstuffs. The method allowed detection of the main antibiotic groups, including tetracyclines. In 2013-2014, a total of 171 feed samples were analysed to determine antibacterial substances. Among the analysed samples 84 (49.1%) were suspected to contain tetracyclines. Out of the 84 feeds analysed using chromatography, 28 (33.3%) contained undeclared tetracyclines, which were identified at concentrations ranging from 0.32 mg kg-1 to 48.98 mg kg-1.