Metabolic profile modifications in milk after enrofloxacin administration studied by liquid chromatography coupled with high resolution mass spectrometry.

High resolution accurate mass spectrometry (HRMS) operating in full scan MS mode was used in the search and identification of metabolites in raw milk from cows medicated with enrofloxacin. Data consisting of m/z features were taken throughout the entire chromatogram of milk samples from medicated animals and were compared with blank samples. Twenty six different compounds were identified. Some of them were attributed to structures related to enrofloxacin while others were dipeptides or tripeptides. Additionally, enrofloxacin was administered in a controlled treatment for three days. Milk was collected daily from the first day of treatment and until four days after in the search for the identified compounds. The obtained data were chemometrically treated by Principal Component Analysis. Samples were classified by this method into three different groups corresponding to days 1-2, day 3 and days 4-7 considering the different concentration profile evolution of metabolites during the days studied. Tentative metabolic pathways were designed to rationalize the presence of the newly identified compounds.

High resolution mass spectrometry in the identification of transformation products and metabolites from ß-lactam antibiotics in thermally treated milk.

Antibiotics such as ß-lactam derivatives (penicillins and cephalosporins) are frequently used in veterinary medicine. The presence of these antibiotics together with their metabolites and/or products produced in subsequent treatments at which milk is submitted (sterilization, pasteurization), may be responsible for bacterial resistance, allergy and/or toxicity on sensitive individuals. In this study, liquid chromatography coupled with high resolution mass spectrometry (LC-HRMS) is used to identify transformation products (TPs) from four ß-lactam antibiotics (amoxicillin (AMOX), cephapirin (PIR), ceftiofur (TIO) and penicillin G (PENG)) in thermally treated cow milk. In addition, milk from cows medicated with PENG has also been analogously treated and studied. The detected TPs come mainly from hydrolysis and decarboxylation reactions. Products more strongly degraded respect to parent compounds (of lower molecular weight) were obtained after treating milk at higher temperatures. Products identified in milk from cows medicated with PENG have been classified as TPs when coming from chemical/thermal degradation, and metabolites when resulting from the biological drug metabolism. While TPs are the result of hydrolysis and decarboxylation processes, as already indicated, an enzymatic conjugation with amino acids is suggested to be involved in the formation of metabolites.

Comparative study of the LC-MS/MS and UPLC-MS/MS for the multi-residue analysis of quinolones, penicillins and cephalosporins in cow milk, and validation according to the regulation 2002/657/EC.

The aim of this study was to develop and validate an analytical method to simultaneously determine European Union-regulated ß-lactams (penicillins and cephalosporins) and quinolones in cow milk. The procedure involves a new solid phase extraction (SPE) to clean-up and pre-concentrate the three series of antibiotics before analysis by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and ultra-high-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). LC-MS/MS and UPLC-MS/MS techniques were also compared. The method was validated according to the Directive 2002/657/EC and subsequently applied to 56 samples of raw cow milk supplied by the Laboratori Interprofessional Lleter de Catalunya (ALLIC) (Laboratori Interprofessional Lleter de Catalunya, Control Laboratory Interprofessional of Milk of Catalunya).